CN105837575A - 3-ethynyl pyrazolo pyrimidine derivative and preparation method and application thereof - Google Patents

3-ethynyl pyrazolo pyrimidine derivative and preparation method and application thereof Download PDF

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CN105837575A
CN105837575A CN201510016197.2A CN201510016197A CN105837575A CN 105837575 A CN105837575 A CN 105837575A CN 201510016197 A CN201510016197 A CN 201510016197A CN 105837575 A CN105837575 A CN 105837575A
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CN105837575B (en
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杨胜勇
魏于全
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Sichuan University
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Sichuan University
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Priority to JP2017554635A priority patent/JP6465996B2/en
Priority to CA2973247A priority patent/CA2973247C/en
Priority to AU2016207167A priority patent/AU2016207167B2/en
Priority to PCT/CN2016/070725 priority patent/WO2016112846A1/en
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5375—1,4-Oxazines, e.g. morpholine
    • A61K31/5377—1,4-Oxazines, e.g. morpholine not condensed and containing further heterocyclic rings, e.g. timolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Abstract

The invention belongs to the field of medicinal chemistry, and specifically, relates to a 3-ethynyl pyrazolo pyrimidine derivative and a preparation method and application thereof. The invention provides the 3-ethynyl pyrazolo pyrimidine derivative having a structural formula I shown in the description. The invention also provides the preparation method and application of the derivative, a medical composition containing the derivative, and an application of the derivative and the medical composition to treatment and prevention of tumors.

Description

3-acetenyl Pyrazolopyrimidine derivative and its production and use
Technical field
The invention belongs to chemical medicine, be specifically related to 3-acetenyl Pyrazolopyrimidine derivative and its production and use.
Background technology
Protein kinase is the phosphotransferase that a class is important, and its function is the γ-phosphate of catalysis adenosine triphosphate (ATP) end Group transfers on the amino acid residue of specific substrates protein, makes protein phosphorylation, thus plays its biochemical functions.Egg White kinases is the key component of intracellular signal transduction pathway.Protein kinase participate in regulate and control multiple cell processes, including cell growth, Energy metabolism, cell cycle, transcribe, apoptosis and differentiation etc..Additionally, protein kinase is maintaining iuntercellular contact, internal flat The aspects such as weighing apparatus, function of immune system also function to vital effect.The exception regulation of protein kinase is the most swollen with multiple disease The generation development of tumor is relevant.Protein kinase has become important disease treatment target, causes and pays close attention to widely.
Since listing to calendar year 2001 first tyrosine protein kinase medicine imatinib (Imatinib), protein kinase medicine becomes For the whole world fastest-rising unit of pharmaceutical market, end 2014 more than 20 protein kinase medicine has been ratified in the world on City, plants anti-tumor medicine, such as imatinib (Imatinib), gefitinib (Gefitinib), strategic point including more than ten Lip river replace Buddhist nun (Erlotinib), Sorafenib (Sorafenib), Sutent (Sunitinib), Dasatinib (Dasatinib), AMN107 (Nilotinib), Lapatinib (Laptinib), pazopanib (Pazopanib) and Rui Gefeini (Regorafenib) Deng.The research and development success of high the research and development success rate, particularly multiple protein kinases anti-tumor drugs targeting of protein kinases medicine, promotees Become the hot research direction of science and pharmaceuticals industry circle.
Although the research and development of current protein kinase medicine achieve huge success, but still there are the biggest development space and potentiality.Especially It is the novel kinases inhibitor of research and development for refractory neoplasm, resistant tumors, the treatment of high metastatic tumo(u)r, such as three Negative breast cancer, drug resistance pulmonary carcinoma, hepatocarcinoma, cancer of pancreas, melanoma, multiple leukemia etc., remain current research heat Point.
Summary of the invention
The invention provides a kind of 3-acetenyl Pyrazolopyrimidine derivative, its structure is as shown in formula I:
Wherein, R1For-H, C1～C4Alkyl,
R2For-H, C1～C8Alkyl,R8Substituted C3～C8Cycloalkyl,Epoxy alkyl,
R3～R7Independently be-H, C1～C8Alkyl ,-OH, C1～C8Alkoxyl, halogen,
R8～R11Independently be-H, C1～C8Alkyl, halogen ,-OH,
R12～R14Independently be
R15～R19Independently be-H, C1～C8Alkyl ,-OH, C1～C8Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, halogen, C1～C8Alkyl, C1～C8Cycloalkyl ,-OCF3Or-CF3；
R39～R42Independently be C1～C8Alkyl, C3～C8Cycloalkyl or C1～C8Hydroxy alkyl；
N=0～6.
As currently preferred technical scheme, R1For-H orR2For-H, C1～C4Alkyl,R8Replace C3～C8Cycloalkyl,C3～C8Epoxy alkyl, R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen, R8～R11Independently be-H, C1～C4Alkyl, halogen ,-OH, R12～R14Independently be R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1~ C4Alkoxyl, halogen ,-CF3、-OCF3、 R20～R38Independently be-H, halogen, C1～C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3；R39～R42Independently For C1～C4Alkyl, C3～C8Cycloalkyl or C1～C4Hydroxy alkyl；N=0～4.
Preferably, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl, R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen, R8～R11Independently be-H, C1～C4Alkyl ,-OH, R12～R14Independently be R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、 -OCF3、R20～R38Independently be-H, halogen Element, C1～C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3；R39～R42Independently be C1～C4Alkyl, C3～C8 Cycloalkyl or C1～C4Hydroxy alkyl；N=0～4.
It is further preferred that R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Ring Alkyl,C3～C8Epoxy alkyl, R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen, R8～R11Independently be-H, C1～C4Alkyl ,-OH, R12～R14Independently be R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, halogen, C1~ C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3；R39～R42Independently be C1～C4Alkyl, C3～C8Cycloalkyl or C1～C4Hydroxy alkyl；N=0～4.
Further preferred, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8 Cycloalkyl,C3～C8Epoxy alkyl, R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen, R8～R11Independently be-H, C1～C4Alkyl ,-OH, R12～R14Independently be R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, halogen, C1～C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3； N=0～2.
Optimum, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl, R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl ,-F ,-Cl, R8～R11Independently be-H, C1～C4Alkyl ,-OH,R12～R14Independent Ground isR15~ R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl ,-F ,-Cl ,-CF3、-OCF3、 R20～R38Independently be-H, C1～C4Alkyl or-CF3；N=0 or 1.
Above-mentioned 3-acetenyl Pyrazolopyrimidine derivative, works as R6ForTime, its structure is as shown in formula II:
Wherein, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl,N=0～4； R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH, R15～R19Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1~ C4Alkyl or-CF3。
Preferably, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl,N=0～4； R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH, R15～R19Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1~ C4Alkyl or-CF3。
Preferably, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8 Epoxy alkyl,N=0～4；R8~ R11Independently be-H, C1～C4Alkyl ,-OH,R3～R5、R7Independently be-H, C1~ C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R12For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, Halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
It is further preferred that R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4 Alkoxy or halogen；R12For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Further preferred, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,N=0～2；R8～R11Independently be -H、C1～C4Alkyl ,-OH,R3～R5、R7Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxy or halogen；R12For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、 -OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Further preferred, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,N=0 or 1；R8～R11Independently be -H、C1～C4Alkyl ,-OH,R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1~ C4Alkoxy or halogen；R12For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl or halogen；R1For-H orR2For-H, C1~ C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,R12For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1~ C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or -CF3。
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl ,-F or-Cl；R1For-H orR2For -H、C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,R12For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1~ C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or -CF3。
Further preferred, R3～R5、R7Independently be-H, methyl or-Cl；R1For-H orR2For-H, C1~ C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,R12For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1~ C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or -CF3。
Preferably, R15～R19Independently be-H, C1～C4Alkyl, C1～C4Alkoxyl, halogen ,-CF3、-OCF3OrR1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl,N=0～4； R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,R12For R20～R38Independently be-H, C1～C4Alkyl Or-CF3。
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, methoxyl group ,-F ,-Cl ,-CF3、-OCF3OrR1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl,N=0～4； R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,R12For R20～R38Independently be-H, C1～C4Alkyl Or-CF3。
Optimum, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl, N=0 or 1；R3～R5、R7Independently be-H, methyl Or-Cl；R8～R11Independently be-H, C1～C4Alkyl ,-OH,R12For R15～R19Independently be-H, C1～C4Alkyl, Methoxyl group ,-F ,-Cl ,-CF3、-OCF3OrR20～R38Independently be-H, C1～C4Alkyl or-CF3。
Above-mentioned 3-acetenyl Pyrazolopyrimidine derivative, works as R6ForTime, its structure is as shown in formula III:
Wherein, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl, N=0～4；R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independent Ground is-H, C1～C4Alkyl ,-OH,R13For R15～R19Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1~ C4Alkyl or-CF3。
Preferably, R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy Alkyl,N=0～4；R8、R9Independently be-H, C1～C4 Alkyl ,-OH,R1For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R13For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, Halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
It is further preferred that R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3~ C8Epoxy alkyl,R8、R9Independently be-H, C1～C4Alkane Base ,-OH,R1For-H orR3～R5、R7Independently be-H, C1~ C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R13For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, Halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl ,-OH, R1For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl or Halogen；R13For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、 R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl,R1For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R13For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Solely It is on the spot-H, C1～C4Alkyl or-CF3。
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or halogen；R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be -H、C1～C4Alkyl ,-OH,R13For R15～R19Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1~ C4Alkyl or-CF3。
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or-Cl；R1For-H orR2 For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be -H、C1～C4Alkyl ,-OH,R13For R15～R19Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1~ C4Alkyl or-CF3。
Preferably, R13ForR1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R3～R5、R7Independently be-H, C1~ C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH, R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Preferably, R15～R19Independently be-H, C1～C4Alkyl, C1～C4Alkoxyl, halogen ,-CF3Or R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3~ C8Epoxy alkyl,N=0～ 4；R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,R13For R20～R38Independently be-H, C1～C4Alkyl Or-CF3。
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, halogen ,-CF3OrR1For-H OrR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,N=0～4；R3~ R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1~ C4Alkyl ,-OH,R13For R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Further preferred, R15～R19Independently be-H, C1～C4Alkyl ,-CF3OrR1For-H or R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R3～R5、R7Independent Ground is-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,R13For R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Preferably, R20～R38Independently be-H or C1～C4Alkyl；R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R3～R5、R7Independently be-H, C1~ C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH, R13For R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
Optimum, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8 Epoxy alkyl,R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or-Cl；R8、R9 Independently be-H, C1～C4Alkyl,R13ForR15~ R19Independently be-H, C1～C4Alkyl ,-CF3OrR20～R38Independently be-H or C1～C4Alkyl.
Above-mentioned 3-acetenyl Pyrazolopyrimidine derivative, works as R6ForR13ForTime, its structure such as formula IV Shown in:
Wherein, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl, N=0～4；R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independent Ground is-H, C1～C4Alkyl ,-OH,R15～R19Independently be-H, C1~ C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
Preferably, R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy Alkyl,N=0～4；R8、R9Independently be-H, C1～C4 Alkyl ,-OH,R1For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1~ C4Alkoxyl, halogen ,-CF3、-OCF3、
It is further preferred that R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3~ C8Epoxy alkyl,R8、R9Independently be-H, C1～C4Alkane Base ,-OH,R1For-H orR3～R5、R7Independently be-H, C1~ C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkane Epoxide, halogen ,-CF3、-OCF3、
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl ,-OH, R1For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl or Halogen；R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl,R1For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R15～R19Independently For-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or halogen；R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be -H、C1～C4Alkyl ,-OH,R15～R19Independently be-H, C1～C4 Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or-Cl；R1For-H orR2 For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be -H、C1～C4Alkyl ,-OH,R15～R19Independently be-H, C1～C4 Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
Preferably, R15～R19Independently be-H, C1～C4Alkyl, C1～C4Alkoxyl, halogen ,-CF3Or R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3~ C8Epoxy alkyl,N=0～ 4；R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, halogen ,-CF3OrR1For-HR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,N=0～4；R3~ R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1~ C4Alkyl ,-OH,Further preferred, R15～R19Independently be-H, C1～C4Alkyl ,-CF3Or
Optimum, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8 Epoxy alkyl,R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or-Cl；R8、R9 Independently be-H, C1～C4Alkyl,R15～R19Independently be-H, C1～C4Alkyl ,-CF3
Above-mentioned 3-acetenyl Pyrazolopyrimidine derivative, works as R6ForR14ForTime, its structure such as formula Shown in V:
Wherein, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl,N=0～4； R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,R15～R19Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1~ C4Alkyl or-CF3。
Preferably, R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl, C3～C8Epoxy alkyl,N=0～4； R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,R15～R19Independently be-H, C1～C4Alkyl, -OH、C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1~ C4Alkyl or-CF3。
Preferably, R2For C1～C4Alkyl,C3～C8Epoxy alkyl, N=0 or 1；R9For C1～C4Alkyl ,-OH,R1 For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R15～R19 Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、 R20～R38Independently be-H, C1～C4Alkyl or-CF3。
It is further preferred that R2For C1～C4Alkyl,R9For C1～C4 Alkyl ,-OH orR1For-H orR3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1~ C4Alkoxy or halogen；R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、 -OCF3、R20～R38Independently be-H, C1～C4Alkyl or-CF3。
Further preferred, R2For C1～C4Alkyl orR9For C1～C4Alkyl；R1For-H or R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Solely It is on the spot-H, C1～C4Alkyl or-CF3。
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or halogen；R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4 Alkyl or-CF3。
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl or halogen；R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4 Alkyl or-CF3。
Further preferred, R3～R5、R7Independently be-H, C1～C4Alkyl or-Cl；R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、R20～R38Independently be-H, C1～C4 Alkyl or-CF3。
Preferably, R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen or-CF3；R1For -H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Ring Oxyalkyl,N=0～4；R3～R5、 R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkane Base ,-OH,
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, halogen or-CF3；R1For-H orR2 For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R3～R5、R7Independently be-H, C1~ C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,
Further preferred, R15～R19Independently be-H or-CF3；R1For-H orR2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；R8～R11Independently be-H, C1～C4Alkyl ,-OH,
Optimum, R1For-H；R2For C1～C4Alkyl orR3～R5、R7Independently be-H, C1～C4Alkane Base or-Cl；R9For C1～C4Alkyl；R15～R19Independently be-H or-CF3。
Above-mentioned 3-acetenyl Pyrazolopyrimidine derivative, its structural formula is:
Present invention also offers the preparation method of above-mentioned 3-acetenyl Pyrazolopyrimidine derivative:
Work as R6ForTime, the synthetic route of compound shown in formula II is:
Work as R6ForR13ForTime, the synthetic route of compound shown in formula III is:
Work as R6ForR13ForTime, the synthetic route of compound shown in formula IV is:
Work as R6ForR14ForTime, shown in formula V, the synthetic route of compound is:
Reaction condition in above-mentioned reaction equation is:
(such as NIS (N-N-iodosuccinimide), NBS, (N-bromo succinyl is sub-at common halogenating agent for a, compound 1 Amine), Br2、I2, ICl, IBr) carry out halogenating reaction and prepare compound 2.
b、R2Alkyl halide (bromine band or iodo) or corresponding sulphonic acid ester (methanesulfonates, p-toluenesulfonic esters, to nitro Benzene sulfonate etc.) in the basic conditions (such as KOH, NaOH, K2CO3、Na2CO3, Cs2CO3, NaH) and compound 2 replacements prepare compound 3.
C, corresponding carboxylated compound under common condensation condition (as condensing agent method, mixed anhydride method, activation are sent out) with The corresponding compound 7～10 connected with amido link being condensed to yield correspondence containing amino-compound.
D, corresponding intermediate is made lower to carry out coupling reaction to the alkynes reagent of one end band blocking group transition metal-catalyzed and obtain corresponding Intermediate, deprotection obtains containing the compound 4 of alkynyl, compound 6, compound 7, compound 10 accordingly the most again And compound 13.
E, corresponding halogen-containing intermediate carry out coupling reaction under transition metal-catalyzed with the intermediate containing alkynyl and obtain formula II, compound shown in III, IV, V, coupling reaction uses palladium catalyst (such as Pd4(PPh3)4、PdAc2、Pd2(dba)3、PdPPh3Cl2 Deng), mantoquita (such as Cu-lyt., protobromide ketone, Hydro-Giene (Water Science). etc.), and suitable organic base or inorganic base (as triethylamine, DIPEA (diisopropylethylamine), potassium carbonate, sodium carbonate, sodium bicarbonate etc.) appropriate solvent (as THF (oxolane), Toluene, DMF (N,N-dimethylformamide), 1,4-dioxane etc.) in obtain in 20～150 DEG C of reactions.
F, compound 12 are esterified intermediate under corresponding alcoholic solution (such as methanol, ethanol etc.), sulphuric acid catalysis accordingly.
G, compound 14 suitable alkali (such as triethylamine, DIPEA etc.) and solvent (as EA (ethyl acetate), THF, Dichloromethane etc.) and at a temperature of (0-50 degree) react with triphosgene and obtain compound 15.
H, compound 15 and compound 6 suitable solvent (as EA (ethyl acetate), THF, dichloromethane, toluene, DMF etc.) in obtain compound 16 in 20～120 DEG C of reactions.
Wherein, R1For-H, C1～C4Alkyl,R2For-H, C1～C8Alkyl,R8Replace C3～C8Cycloalkyl,C3～C8Epoxy alkyl, R3～R7Independently be-H, C1～C8Alkyl ,-OH, C1～C8Alkoxyl, halogen, R8～R11Independently be-H, C1～C8Alkyl, halogen ,-OH, R12～R14Independently be R15～R19Independently be-H, C1～C8Alkyl ,-OH, C1~ C8Alkoxyl, halogen ,-CF3、-OCF3、 R20～R38Independently be-H, halogen, C1～C8Alkyl, C1～C8Cycloalkyl ,-OCF3Or-CF3；R39～R42Independently For C1～C8Alkyl, C3～C8Cycloalkyl or C1～C8Hydroxy alkyl；N=0～6.
The 3-acetenyl Pyrazolopyrimidine derivative that the present invention is above-mentioned, includes their isotopic compound, racemic modification, rotation Optical activity isomer, polymorphic forms or its mixture.
Present invention also offers above-mentioned 3-acetenyl Pyrazolopyrimidine derivative pharmaceutically acceptable salt.
Present invention also offers the prodrug of the compounds of this invention, according to the present invention, prodrug is the derivant of above-claimed cpd, they Self be likely to be of more weak activity or even without activity, but upon administration, in physiological conditions (such as by metabolism, Solvolysis or other mode) it is converted to corresponding biologically active form.
Present invention also offers the above-mentioned 3-pharmaceutically acceptable hydrate of acetenyl Pyrazolopyrimidine derivative.
The present invention also provides for a kind of pharmaceutical composition, is that above-mentioned 3-acetenyl Pyrazolopyrimidine derivative provided by the present invention adds Pharmaceutically the complementary composition of acceptable is prepared from.The 3-acetenyl Pyrazolopyrimidine derivative structure that the present invention provides is such as Shown in formula I～V.
Present invention also offers above-mentioned 3-acetenyl Pyrazolopyrimidine derivative, its salt or hydrate in preparing inhibitors of kinases Purposes.
Further, above-mentioned inhibitors of kinases is suppression SRC family TYR protein kinase (Blk TYR protein kinase, Fgr TYR protein kinase, Frk TYR protein kinase, Fyn TYR protein kinase, Hck TYR protein kinase, Lck network Propylhomoserin protein kinase, Lyn TYR protein kinase, c-SRC TYR protein kinase, YES tyrosine protein kinase), FLT3 (people's FMS sample tyrosine kinase 3), Abl (Abl TYR protein kinase), VEGFR1 (vascular endothelial growth factor receptor 1), VEGFR2 (VEGF R2), VEGFR3 (VEGF R3), RET (RET Receptor tyrosine kinase), c-RAF (c-RAF serine/threonine protein kitase), B-RAF (B-RAF serine/threonine Protein kinase), c-KIT (tyrosine protein kinase KIT), PDGF α (platelet derived growth factor receptor), PDGF β (blood Platelet derived growth factor receptor β), FGFR1 (fibroblast growth factor acceptor 1), FGFR2 (Desmocyte growth factor Sub-receptor 2), FGFR3 (fibroblast growth factor receptor3), EphA2 (EphA2 tyrosine kinase), EphB2 (EphB2 Tyrosine kinase), EphB4 (EphB4 tyrosine kinase), ALK (anaplastic lymphoma kinase ALK Alk receptor tyrosine kinase), Met (Met tyrosine kinase) DDR1 (DDR1 TYR kinases), DDR2 (DDR2 TYR kinases), Btk (Btk TYR kinases), BMX (BMX TYR kinases), at least one kinase whose medicine in TAK1 (transforming growth factor kinases 1).(please revise according to practical situation Or delete above-mentioned kinase whose kind)
Present invention also offers above-mentioned 3-acetenyl Pyrazolopyrimidine derivative, its salt or hydrate in preparing antitumor drug Purposes.
Further, above-mentioned tumor is leukemia or solid tumor.
Further, above-mentioned entity tumor is pulmonary carcinoma, breast carcinoma, cancer of pancreas, melanoma, glioma, hepatocarcinoma, first At least one in shape adenoma, cervical cancer, gastric cancer or colorectal cancer.Wherein, above-mentioned leukemia be acute myeloid leukaemia or Mixed type leukemia.
The 3-acetenyl Pyrazolopyrimidine derivative that the present invention provides is to people's pulmonary carcinoma, human breast carcinoma, human pancreas cancer, people's maligna The element tumor such as tumor, human leukemia all shows good inhibition.
Accompanying drawing explanation
Fig. 1 compound 3 internal pharmacodynamic experiment to nude mouse.
Fig. 2 compound 31 internal pharmacodynamic experiment to nude mouse.
Fig. 3 compound 31 vascular study situation to FLK1 transgenic zebrafish under variable concentrations.
Detailed description of the invention
The preparation of embodiment 13-acetenyl-1-isopropyl-1H-pyrazolo [3,4-d] pyrimidine-4-amine (intermediate 4-1)
The first step, preparation 3-iodo-1H-pyrazolo [3,4-d] pyrimidine-4-amine:
1H-pyrazolo [3,4-d] pyrimidine-4-amine (20g, 148.0mmol, 1.0eq), in there-necked flask, adds 150mL DMF (N, N- Dimethylformamide), stirring nitrogen are replaced 3 times, add NIS (N-N-iodosuccinimide) (50g, 222.0mmol, 1.5eq) After in 80 DEG C of reactions, TLC monitors, and reacts complete after 22h.Stopped reaction, concentrating under reduced pressure DMF, to surplus half, adds full And Na2O3S2Aqueous solution 150mL stirs, and filtration under diminished pressure, saturated Na used successively by filter cake2O3S2Aqueous solution and water wash to filtrate Colourless, vacuum drying obtains target product, pale yellow powder (33.9g, yield 87.8%).1H NMR(400MHz,DMSO-d6) δ8.17(s,1H)。MS m/z(ESI):262.1[M+H]。
Second step, preparation 3-iodo-1-isopropyl-1H-pyrazolo [3,4-d] pyrimidine-4-amine:
3-iodo-1H-pyrazolo [3,4-d] pyrimidine-4-amine (5g, 19.2mmol, 1.0eq) in there-necked flask, add 40mL DMF and After potassium carbonate (5.3g, 38.4mmol, 2.0eq), replace 3 times with nitrogen, add 2-N-Propyl Bromide (1.9mL, 20.1mmol, 1.05eq) After in 80 DEG C of reactions, TLC monitors, and reacts complete after 4h.Stopped reaction, decompression boils off DMF, with DCM (dichloromethane) And water extracts 3 times, it is evaporated after merging DCM layer, is EA (ethyl acetate) PE (petroleum ether)=1 3 by volume ratio Mixed solvent recrystallization obtains target product, and in recrystallization mother liquor, target product can obtain through column chromatography for separation, pale yellow powder (5.4g, Yield 92.8%).1H NMR(400MHz,DMSO-d6) δ 8.19 (s, 1H), 4.99-4.93 (m, 1H), 1.42 (d, J=6.7 Hz,6H)。MS m/z(ESI):304.0[M+H]。
3rd step, preparation 1-isopropyl-3-((trimethyl silyl) acetenyl)-1H-pyrazolo [3,4-d] pyrimidine-4-amine:
3-iodo-1-isopropyl-1H-pyrazolo [3,4-d] pyrimidine-4-amine (5.4g, 17.8mmol, 1.0eq), in there-necked flask, adds DMF 40mL,CuI(339mg,1.78mmol,0.1eq),Pd(PPh3)4(1g, 0.89mmol, 0.05eq), nitrogen adds three after replacing 3 times Ethamine (5mL, 35.6mmol, 2.0eq), trimethylsilyl acetylene (2.65mL, 18.7mmol, 1.05eq), react 2.5h in 80 DEG C, TLC monitoring decompression after completion of the reaction boils off DMF, and remaining residue obtains target product through column chromatography for separation, is directly used in lower step anti- Should.MS m/z(ESI):274.2[M+H].
4th step, preparation 3-acetenyl-1-isopropyl-1H-pyrazolo [3,4-d] pyrimidine-4-amine:
1-isopropyl-3-((trimethyl silyl) acetenyl)-1H-pyrazolo [3,4-d] pyrimidine-4-amine is in 20mL MeOH (methanol) In, it is complete that addition potassium carbonate (4.9g, 35.6mmol, 2.0eq) is stirred at room temperature 10min, TLC monitoring reaction, and decompression boils off MeOH Afterwards with water-dispersible, DCM washing water layer 3 times, merge DCM layer, after being evaporated, obtain intermediate 4-1, light ash through column chromatography for separation Color powder (1.6g, yield 44.7%).1H NMR(400MHz,DMSO-d6)δ8.22(s,1H),5.02–4.98(m,1H), 4.62 (s, 1H), 1.44 (d, J=6.7Hz, 6H).MS m/z(ESI):202.1[M+H].
The preparation of embodiment 2～10 intermediate 4-2～4-10
Use different halogenated alkane to react with 1H-pyrazolo [3,4-d] pyrimidine-4-amine, add cesium carbonate, potassium carbonate, DIPEA (two Wopropyl ethyl amine) or other inorganic or organic bases, PdCl2(PPh3)2Or Pd (PPh3)4Deng raw material, by with prepare intermediate 1 Described similar method obtains following intermediate 4-2～4-10.
The preparation of the iodo-1-of embodiment 113-(1-(methylsulfonyl) piperidines-3-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine (intermediate 4-11)
The first step, prepares the tert-butyl group-3-((methylsulfonyl) oxo) piperidines-1-carboxylate:
N-Boc-3-hydroxy piperidine (2.1g, 10mmol, 1.0eq), DIPEA (2.1mL, 15mmol, 1.5eq) is in 20mL DCM In, stirring, it is slowly added dropwise mesyl chloride (1.0mL, 13mmol, 1.3eq) after being cooled to 0 DEG C and finishes rear natural reaction to room temperature. Then reactant liquor uses 1MHCl solution, saturated sodium bicarbonate aqueous solution, saturated sodium-chloride water solution and water washing, DCM successively Layer is dried through anhydrous magnesium sulfate, is evaporated to obtain product, yellow solid (2.65g, yield 95.1%) after filtration.MS m/z(ESI):280.1 [M+H]。
Second step, prepares the tert-butyl group-3-(4-amino-3-iodo-1H-pyrazolo [3,4-d] pyrimidine-1-base) piperidines-1-carboxylate:
3-iodo-1H-pyrazolo [3,4-d] pyrimidine-4-amine (2.4g, 9.2mmol, 1.0eq), the tert-butyl group-3-((methylsulfonyl) oxo) piperidines-1- Carboxylate (3.1g, 11mmol, 1.2eq), cesium carbonate (6.0g, 18.4mmol, 2.0eq) is in 50mL DMF, and nitrogen is replaced 80 DEG C of reactions after 3 times, react complete after TLC monitoring 3h.Being evaporated DMF, water-dispersible, DCM is extracted to water layer without product, Merge DCM layer, after being evaporated, obtain target product, faint yellow solid (2.9g, yield 73.4%) through column chromatography for separation.1H NMR (400MHz,CDCl3)δ8.31(s,1H),6.12(s,2H),4.78-4.70(m,1H),4.31(br.s,1H),4.12(br.s,1H), 3.36 (br.s, 1H), 2.83 (t, J=12.0Hz, 1H), 2.30-1.94 (m, 2H), 1.87 (d, J=13.0Hz, 1H), 1.74-1.54 (m,1H),1.44(s,9H)。MS m/z(ESI):445.1[M+H]。
3rd step, the preparation iodo-1-of 3-(piperidines-3-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine:
The tert-butyl group-3-(4-amino-3-iodo-1H-pyrazolo [3,4-d] pyrimidine-1-base) piperidines-1-carboxylate (2.9g) is in methanol 35mL Dissolve, add 4MHCl dioxane solution 35mL, separate out solid after 8h is stirred at room temperature, then reactant liquor is cooled to 0 DEG C Rear filtration under diminished pressure, filter cake is dispersed in water, and separates out solid, filtration under diminished pressure, wash solid after adjusting PH to 8, and solid is in ethanol In, decompression boils off ethanol final vacuum and is dried to obtain target product (2.2g, yield 93.1%).MS m/z(ESI):345.1[M+H].
4th step, the preparation iodo-1-of 3-(1-(methylsulfonyl) piperidines-3-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine:
The iodo-1-of 3-(piperidines-3-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine (172.1mg, 0.5mmol, 1.0eq) is in 4mL DCM In, add after DIPEA (129.24mg, 1.0mmol, 2.0eq) in 0 DEG C of stirring, be slowly added dropwise mesyl chloride (57.3mg, 0.5mmol, 1.0eq), finishing rear TLC, confirm that reactant liquor uses saturated sodium bicarbonate aqueous solution successively after completion of the reaction, saturated sodium-chloride is water-soluble Liquid and water washing, DCM layer is dried through anhydrous magnesium sulfate, is evaporated to obtain intermediate 4-11 after filtration, white powder (197.8mg, Yield 93.7%).1H NMR(400MHz,DMSO-d6) δ 8.23 (s, 1H), 4.83-4.65 (m, 1H), 3.69 (d, J=7.2 Hz, 1H), 3.59 (d, J=11.1Hz, 1H), 3.07 (t, J=10.9Hz, 1H), 2.91 (s, 3H), 2.80 (t, J=10.9Hz, 1H), 2.45-2.00 (m, 2H), 1.95 (d, J=13.6Hz, 1H), 1.76-1.59 (m, 1H).MS m/z(ESI):423.0[M+H].
The preparation of embodiment 12～19 intermediate 4-12～4-19
React with 1H-pyrazolo [3,4-d] pyrimidine-4-amine with different alkylols or alkyl sulfonic ester, add cesium carbonate, potassium carbonate, Other inorganic or organic bases, mesyl chloride or to nitro mesyl chloride, HCl ethanol solution, HCl diethyl ether solution, trifluoro second The raw materials such as acid, by with prepare method similar described in intermediate 4-11 and obtain following intermediate 4-12～4-19.
The preparation of the iodo-1-of embodiment 20 (R)-3-(1-methyl piperidine-3-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine (intermediate 4-20)
(R) the iodo-1-of-3-(piperidines-3-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine (750mg, 2mmol, 1.0eq) is dissolved in 18mL 1,2- In the mixed solvent of dichloroethanes methanol=8 1 (volume ratio), be slowly added dropwise formalin (37%, 0.82mL, 10mmol, 5.0eq), 10min is stirred at room temperature after finishing, adds NaBH in two batches3CN (502.7mg, 8mmol, 4.0eq), after 10min TLC monitoring reaction is complete, drips water 3mL, finish rear evaporated under reduced pressure solvent in reactant liquor, and residue is dispersed in water, and adjusts PH, to alkalescence, filters, and washes solid, then solid evaporated under reduced pressure in ethanol, and residue obtains intermediate 4-20 through recrystallization, White powder (500mg, yield 70.0%).1H NMR(400MHz,DMSO-d6)δ8.20(s,1H),4.73–4.66(m, 1H), 4.41 (t, J=4.8Hz, 1H), 3.48-3.41 (m, 1H), 2.88 (dd, J=11.1,3.7Hz, 1H), 2.77 (d, J=11.1 Hz,1H),2.21(s,3H),1.97-1.72(m,3H),1.71-1.61(m,1H)。MS m/z(ESI):359.1[M+H]。
The preparation of embodiment 21～24 intermediate 4-21～4-24
React with 1H-pyrazolo [3,4-d] pyrimidine-4-amine with the secondary amine of different band aryl, add the raw material such as alkyl aldehydes or aryl aldehyde, By with prepare method similar described in intermediate 4-20 and obtain following intermediate 4-21～4-24.
Embodiment 25 (R)-1-((R)-3-(4-amino-3-iodo-1H-pyrazolo [3,4-d] pyrimidine-1-base) piperidin-1-yl)-2-hydroxy propane-1-ketone The preparation of (intermediate 4-25)
By D-2-hydracrylic acid (157mg, 1.74mmol, 1.2eq), HOBT (I-hydroxybenzotriazole) (235.1mg, 1.74mmol, 1.2eq), EDCI (1-ethyl-3-(3-dimethylamine propyl) carbodiimide hydrochloride) (417mg, 2.2mmol, 1.5eq) It is dissolved in DMF, adds triethylamine (0.61mL, 4.35mmol, 3.0eq), after 0.5h is stirred at room temperature, add (R)-3-iodo-1-(piperazine Pyridine-3-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine (500mg, 1.45mmol, 1.0eq) room temperature reaction about 5h, after be evaporated DMF, extracts 2 times with dichloromethane with saturated sodium bicarbonate aqueous solution, is evaporated after merging organic layer, and residue divides through column chromatography From obtaining intermediate 4-25, white solid (442mg, yield 73.3%).
1H NMR(400MHz,DMSO-d6)δ8.22(s,1H),5.04–4.94(m,1H),4.77-4.51(m,1H),4.50 4.44 (m, 1H), 4.40-4.26 (m, 1H), 4.19-4.03 (m, 1H), 3.13 2.98 (m, 1H), 2.76 (t, J=12.1Hz, 1H), 2.25-2.11 (m, 1H), 2.06 2.04 (m, 1H), 1.86 (t, J=13.8Hz, 1H), 1.62 1.49 (m, 1H), 1.24 1.13(m,3H)。MS m/z(ESI):417.1[M+H]。
The preparation of embodiment 26-31 intermediate 4-26～4-31
With raw materials such as the secondary amine of different band aryl, the lactic acid of various configuration, use and prepare method similar described in intermediate 4-25 Obtain following intermediate 4-26～4-31.
The preparation of embodiment 324-(4-amino-3-iodo-1H-pyrazolo [3,4-d] pyrimidine-1-base) hexamethylene-1-alcohol (intermediate 4-32)
The first step: with reference to the method described in embodiment 11 first step, with 4-hydroxy-cyclohexanone Glycol Acetal and methylsufonyl chloride be Material synthesis obtains target product.MS m/z(ESI):236.1[M+H].
Second step, the preparation iodo-1-of 3-(Isosorbide-5-Nitrae-dioxo spiro [4.5] decane-8-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine:
With reference to method described in embodiment 11 second step with 1,4-dioxo spiro [4.5] decane-8-base methanesulfonates and 3-iodo-1H-pyrazolo [3,4-d] pyrimidine-4-amine is that Material synthesis obtains target product.1H NMR(400MHz,DMSO-d6)δ8.19(s,1H),4.79- 4.63(m,1H),3.98-3.82(m,4H),2.19-2.11(m,2H),1.91-1.64(m,6H)。MS m/z(ESI):402.2 [M+H]。
3rd step, preparation 4-(4-amino-3-iodo-1H-pyrazolo [3,4-d] pyrimidine-1-base) hexamethylene-1-ketone:
The iodo-1-of 3-(1,4-dioxo spiro [4.5] decane-8-base)-1H-pyrazolo [3,4-d] pyrimidine-4-amine (1.94g, 4.84mmol) in In 40mL acetone, add 1MHCl 20mL, after 70 DEG C of 3h, be down to room temperature reaction overnight, after decompression boil off acetone, Remaining aqueous solution adjusts pH to 10, separates out a large amount of solid, filters, and is dried to obtain target product, white solid (1.68g, yield 97.1%).1H NMR(400MHz,DMSO-d6)δ8.27(s,1H),5.21–5.15(m,1H),2.75-2.61(m,2H),2.37– 2.26(m,4H),2.20–2.17(m,2H)。MS m/z(ESI):358.1[M+H]。
4th step, preparation 4-(4-amino-3-iodo-1H-pyrazolo [3,4-d] pyrimidine-1-base) hexamethylene-1-alcohol:
4-(4-amino-3-iodo-1H-pyrazolo [3,4-d] pyrimidine-1-base) hexamethylene-1-ketone (1.68g, 4.7mmol, 1.0eq) in methanol, It is dividedly in some parts sodium borohydride (183mg, 4.7mmol, 1.0eq) after being cooled to 0 DEG C, after finishing, is naturally warmed to room temperature reaction, 1h Rear reaction is complete, adds solvent evaporated after 5mL water stirring 5min, and residue is through dichloromethane and saturated sodium bicarbonate aqueous solution After extraction, column chromatography obtains intermediate 4-32, white solid (844mg, yield 50.1%).1H NMR(400MHz,DMSO-d6) δ 8.19 (s, 1H), 4.68 (d, J=4.3Hz, 1H), 4.62-4.50 (m, 1H), 3.56-3.49 (m, 1H), 2.03-1.79 (m, 5H), 1.46-1.29(m,3H)。MS m/z(ESI):360.2[M+H]。
Embodiment 33N-(3-iodo-4-aminomethyl phenyl)-4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) benzamide (intermediate 33) Preparation
The first step, preparation 4-methyl-3-Trifluoromethyl-benzoic acid methyl ester:
4-methyl-3-(trifluoromethyl) benzoic acid (2.04g, 10mmol, 1.0eq), in 25mL MeOH, adds 2mL under stirring Concentrated sulphuric acid, back flow reaction, TLC monitors, reacts complete after 24h hour, is evaporated MeOH, DCM and dissolves, successively with saturated Sodium bicarbonate aqueous solution, saturated sodium-chloride water solution and water washing, DCM layer is dried through anhydrous magnesium sulfate, is evaporated after filtration Product (1.85g, yield 84.8%) is used for lower step.
Second step, preparation 4-bromomethyl-3-Trifluoromethyl-benzoic acid methyl ester:
4-methyl-3-(trifluoromethyl) essence of Niobe is dissolved in 1, in 2-dichloroethanes (DCE), adds NBS (N-bromine under stirring For succimide) (1.81g, 10.18mmol, 1.2eq), AIBN (azodiisobutyronitrile) (0.139g, 0.848mmol, 0.1eq) Rear nitrogen is replaced 3 times, and after 80 DEG C of reaction 30h, reactant liquor is successively with saturated sodium bicarbonate aqueous solution, saturated sodium-chloride water solution And water washing, after DCE layer is dried over magnesium sulfate, be evaporated product is directly used in lower step, based on yield 80%.
3rd step, preparation 4-(4-methylpiperazine-1-yl methyl)-3-Trifluoromethyl-benzoic acid methyl ester:
4-bromomethyl-3-trifluoromethyl benzoic acid methyl ester (6.78mmol, 1.0eq), triethylamine (1.03g, 10.2mmol, 1.5eq), N-methyl piperazine (0.681g, 6.8mmol, 1.0eq) in chloroform, 80 DEG C of reactions, react complete after 1h.Reactant liquor is used successively Saturated sodium bicarbonate aqueous solution, saturated sodium-chloride water solution and water washing, chloroform layer is evaporated rear residue and obtains mesh through column chromatography for separation Mark product, pale yellow oil (1.72g, yield 79.9%).MS m/z(ESI):317.2[M+H].
4th step, preparation 4-(4-methylpiperazine-1-yl methyl)-3-Trifluoromethyl-benzoic acid:
4-(4-methylpiperazine-1-yl methyl)-3-Trifluoromethyl-benzoic acid methyl ester (1.72g, 5.4mmol), in 20ml ethanol, adds Entering 5M NaOH aqueous solution 3ml, pH to 6 after stirred overnight at room temperature, concentrating under reduced pressure, concentrated solution reduces pressure after adding oxolane Filtering, filtrate is evaporated to obtain target product, pale yellow powder (1.47g, yield 89.9%).
5th step, preparation N-(3-iodo-4-aminomethyl phenyl)-4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) benzamide:
4-(4-methylpiperazine-1-yl methyl)-3-Trifluoromethyl-benzoic acid (0.755g, 2.5mmol, 1.0eq), HATU (O-(7- Azo BTA-1-oxygen)-N, N, N ', N '-tetramethylurea hexafluorophosphate) (1.14g, 3mmol, 1.2eq), DIPEA (1.29g, 10mmol, 4.0eq) is dissolved in dichloromethane and 0.5h is stirred at room temperature, addition 3-iodo-4-monomethylaniline. (582.6mg, 2.5mmol, 1.0eq) after in 45 DEG C reaction overnight, be evaporated reactant liquor, saturated sodium bicarbonate aqueous solution and dichloro after completion of the reaction Methane extracts three times, merges concentrating under reduced pressure after organic layer, and residue obtains intermediate 33, faint yellow solid through column chromatography for separation (892mg, yield 69.0%).1H NMR(400MHz,CDCl3) 8.10 (s, 1H), 8.08 (d, J=2Hz, 1H), 8.08- 8.05 (m, 1H), 8.0 (s, 1H), 7.95 (s, 1H), 7.55 (dd, J=4,2Hz, 1H), 7.20 (d, J=8Hz, 1H), 3.75 (s, 2H) 2.60-2.41(m,8H),2.40(s,3H),2.30(s,3H)。MS m/z(ESI):518.1[M+H]。
Embodiment 34N-(3-iodo-4-aminomethyl phenyl)-4-((4-methylpiperazine-1-yl) methyl) benzamide (intermediate 34)
Intermediate 34 is obtained according to the method for condensing that the 5th step described in intermediate 33 is similar, faint yellow solid (532mg, hydrochlorate, Yield 76.7%).1H NMR(400MHz,DMSO-d6)δ10.35(br.s,1H),10.26(s,1H),8.34(s,1H),7.95 (d, J=7.6Hz, 2H), 7.72 (d, J=8.3Hz, 1H), 7.47 (d, J=7.6Hz, 2H), 7.30 (d, J=8.3Hz, 1H), 3.64 (s, 2H),3.39(br.s,2H),3.01(br.s,2H),2.87(br.s,2H),2.73(s,3H),2.42(br.s,2H),2.34(s,3H)。MS m/z(ESI):450.1[M+H]。
Embodiment 35N-(3-acetenyl-4-aminomethyl phenyl)-4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) benzamide is (middle Body 35) preparation
The first step, preparation 3-trimethylsilyl acetylene base-4-monomethylaniline.:
By iodo-for 3-4-monomethylaniline. (11.65g, 50mmol, 1.0eq), Hydro-Giene (Water Science). (0.9g, 4.7mmol, 0.1eq), double Triphenylphosphine palladium (1.75g, 2.5mmol, 0.05eq) is dissolved in dioxane, and nitrogen is replaced three times, adds DIPEA (1.29g, 100mmol, 2.0eq), trimethylsilyl acetylene (6.4g, 65mmol, 1.3eq).It is warming up to 75 DEG C of reactions, Reacting complete after 17h, evaporated under reduced pressure solvent, residue obtains target product through column chromatography for separation, is directly used in the next step.
Second step, preparation 3-acetenyl-4-monomethylaniline.:
3-trimethylsilyl acetylene base-4-monomethylaniline. methanol dissolves, and adds potassium carbonate (3.45g, 25mmol, 0.5eq), room temperature React complete after stirring 10min.Evaporated under reduced pressure solvent, residue DCM, water extraction 3 times, DCM layer is evaporated after drying, Obtain grease, obtain product 3-acetenyl-4-monomethylaniline., yellowish-brown grease (4.0g, yield 61.0%) through column chromatography for separation.1H NMR(400MHz,CDCl3) δ 6.99 (d, J=8.1Hz, 1H), 6.81 (s, 1H), 6.61 (d, J=8.1Hz, 1H), 3.44 (s,2H),3.23(s,1H),2.34(s,3H)。MS m/z(ESI):132.2[M+H]。
3rd step, preparation N-(3-acetenyl-4-aminomethyl phenyl)-4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) benzamide:
Intermediate 35 is obtained according to method of condensing similar described in intermediate 33 the 5th step, faint yellow solid (1.2g, hydrochlorate, Yield 71.7%).1H NMR(400MHz,DMSO-d6) δ 11.24 (br.s, 1H), 10.58 (s, 1H), 8.33 (d, J=6.4Hz, 2H), 8.14 (s, 1H), 7.92 (s, 1H), 7.72 (d, J=8.3Hz, 1H), 7.28 (d, J=8.3Hz, 1H), 4.39 (s, 1H), 4.06 (s, 2H), 3.47 (d, J=10.9Hz, 2H), 3.22 (br.s, 2H), 3.13 (br.s, 2H), 2.89 (br.s, 2H), 2.77 (s, 3H), 2.36 (s,3H)。MS m/z(ESI):416.3[M+H]。
The preparation of embodiment 36N-(3-acetenyl-4-aminomethyl phenyl)-6-(trifluoromethyl) picoline amide (intermediate 36)
6-trifluoromethylpyridin-2-formic acid (229.33mg, 1.2mmol, 1.2eq) thionyl chloride is dissolved, 80 DEG C of backflow 3h, Evaporated under reduced pressure thionyl chloride, obtains 6-(trifluoromethyl) pyridine-2-formyl chloride, colorless oil, dissolves with DCM.By 3-acetylene DCM used by base-4-monomethylaniline. (131.17mg, 1.0mmol, 1.0eq) and triethylamine (202.4mg, 2.0mmol, 2.0eq) Dissolve, be cooled to-5 DEG C, 6-trifluoromethylpyridin-2-formyl chloride DCM solution is slowly dropped to wherein.After dropping, Warm naturally to room temperature, after 15min, react complete.Adding sodium hydrate aqueous solution in reactant liquor, DCM extracts 2 times.Close And DCM layer, then extract with dilute hydrochloric acid, water successively, DCM layer anhydrous magnesium sulfate is dried, and is evaporated to obtain intermediate 36, white Solid (225mg, yield 73.9%).1H NMR(400MHz,DMSO-d6)δ10.47(s,1H),8.08-8.05(m,2H), 7.93 (dd, J=7.3,1.4Hz, 1H), 7.76 (d, J=2.2Hz, 1H), 7.54 (dd, J=8.3,2.2Hz, 1H), 7.06 (d, J=8.3 Hz,1H),4.16(s,1H),2.13(s,3H)。MS m/z(ESI):305.1[M+H]。
The preparation of embodiment 37-61 intermediate 37-61
With the aryl formate containing different substituents, aryl-acyl chlorides, five-ring heterocycles acyl chlorides and the aniline containing different substituents as raw material, Use and prepare method similar described in intermediate 36 and obtain following intermediate 37-61.
The preparation of embodiment 622-tert-butyl-n-(3-acetenyl-4-aminomethyl phenyl) thiazole-5-Methanamide (intermediate 62)
The first step, preparation 2-tert-Butyl-thiazol-5-carboxylic acid, ethyl ester:
By 3-BrPA ethyl ester (1.95g, 10mmol, 1.0eq) and 2,2,2-trimethylthioacetamide (1.17g, 10mmol, 1.0eq) join in 20mL ethanol, stirring at normal temperature 48h.After completion of the reaction, evaporated under reduced pressure solvent, residue bicarbonate Sodium water solution extracts with DCM, and DCM layer anhydrous magnesium sulfate is evaporated after drying, and residue obtains the tertiary fourth of 2-through column chromatography for separation Base-thiazole-5-carboxylic acid ethyl ester, colorless oil (1.15g, yield 55.0%).1H NMR(400MHz,CDCl3)δ8.01(s, 1H), 4.38 (q, J=7.1Hz, 2H), 1.46 (s, 9H), 1.37 (t, J=7.1Hz, 3H).MS m/z(ESI):214.0[M+H].
Second step, preparation 2-tert-Butyl-thiazol-5-carboxylic acid:
2-tert-Butyl-thiazol-5-carboxylic acid, ethyl ester is dissolved in 10mL THF, adds Lithium hydrate (0.45g, 10.9mmol, 1.1eq) Aqueous solution 5mL, stirring at normal temperature about 16h.After completion of the reaction, evaporated under reduced pressure solvent, residue 5mL water dissolution, use 2M HCl Regulation pH to about 3, separates out a large amount of white fluffy solid, sucking filtration, washs filter cake with water (2 × 5mL), obtain the tertiary fourth of 2- Base-thiazole-5-carboxylic acid, white solid (0.55g, yield 55.0%).MS m/z(ESI):184.0[M-H].
3rd step, preparation 2-tert-Butyl-thiazol-5-carbonyl chlorination:
2-tert-Butyl-thiazol-5-carboxylic acid (203.7mg, 1.1mmol) is dissolved in 3ml thionyl chloride, subtracts after 80 DEG C of backflow 2h Pressure solvent evaporated, is directly used in next step.
4th step, preparation 2-tert-butyl-n-(3-acetenyl-4-aminomethyl phenyl) thiazole-5-Methanamide:
3-acetenyl 4-metlyl-phenylamine (131.2mg, 1.0mmol, 1.0eq) is dissolved in 4mL DCM, add triethylamine (151.8mg, 1.5mmol, 1.5eq), it is slowly added dropwise the dichloromethane solution of step 2-tert-Butyl-thiazol-5-carbonyl chlorination after being cooled to 0 DEG C (4mL), reaction 10min, the most successively with saturated aqueous ammonium chloride, saturated sodium bicarbonate water naturally it are warmed to room temperature after finishing Solution, saturated sodium-chloride water solution washing reaction liquid, organic layer filters after drying through anhydrous magnesium sulfate, evaporated under reduced pressure solvent, 3mL Target product, pale yellow powder (253.6mg, yield 85.1%) is obtained after petroleum ether residue.1H NMR(400MHz, DMSO-d6) δ 9.95 (s, 1H), 8.30 (s, 1H), 7.95 (d, J=2.0Hz, 1H), 7.76 (dd, J=8.3,2.0Hz, 1H), 7.28 (d, J=8.3Hz, 1H), 4.39 (s, 1H), 2.37 (s, 3H), 1.47 (s, 9H).MS m/z(ESI):299.1[M+H].
The preparation of embodiment 63N-(4-chloro-3-ethynyl phenyl)-3-(trifluoromethyl) benzamide (intermediate 63)
First and second step, preparation 3-acetenyl-4-chloroaniline:
With 3-iodo-4-chloroaniline as raw material, according to the method described in intermediate 35 first and second step, obtain target product, faint yellow Grease (2.1g, yield 77.8%).1H NMR(400MHz,DMSO-d6) δ 7.12 (d, J=8.7Hz, 1H), 6.75 (d, J =2.8Hz, 1H), 6.60 (dd, J=8.7,2.8Hz, 1H), 5.39 (s, 2H), 4.34 (s, 1H).MS m/z(ESI):518.3[M+H].
3rd step, preparation N-(4-chloro-3-ethynyl phenyl)-3-(trifluoromethyl) benzamide:
The similar approach described in intermediate 36 is used to obtain intermediate 63, pale yellow powder (291mg, yield 90.1%).1H NMR (400MHz,DMSO-d6) δ 10.64 (s, 1H), 8.30 (s, 1H), 8.26 (d, J=8.0Hz, 1H), 8.07 (d, J=2.5Hz, 1H), 8.00 (d, J=8.0Hz, 1H), 7.89-7.82 (m, 1H), 7.82-7.75 (m, 1H), 7.57 (d, J=8.8Hz, 1H), 4.61 (s, 1H)。MS m/z(ESI):324.1[M+H]。
Embodiment 643-iodo-4-methyl-N-(4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) phenyl) benzamide (intermediate 64) Preparation
The first step, preparation 1-bromomethyl-4-nitro-2-trifluoromethyl-benzene:
1-methyl-4-nitro-2-trifluoromethyl-benzene (5.62g, 27.4mmol, 1.0eq) is dissolved in 50ml 1,2-dichloroethanes, so After be sequentially added into N-bromo-succinimide (5.85g, 32.8mmol, 1.2eq) and AIBN (450mg, 2.7mmol, 0.1eq), Heated overnight at reflux.Then reactant liquor is cooled to room temperature, successively with saturated sodium bicarbonate solution, saturated nacl aqueous solution and water Washing, anhydrous magnesium sulfate is dried, and after filtration, concentrating under reduced pressure obtains target product, is directly used in the next step.
Second step, preparation 1-methyl-4-(4-nitro-2-(trifluoromethyl) benzyl) piperazine:
Upper step 1-bromomethyl-4-nitro-2-trifluoromethyl-benzene (20.55mmol) is dissolved in 50mL DCM, adds triethylamine (3.1g, 30.8mmol, 1.5eq) and N methyl piperazine (4.12g, 41.1mmol, 2.0eq), room temperature reaction, react complete Rear concentrating under reduced pressure, DCM dissolves, successively with saturated sodium bicarbonate solution, saturated nacl aqueous solution and water washing, anhydrous magnesium sulfate Being dried, concentrating under reduced pressure after filtration, residue obtains target product, yellow solid through column chromatography for separation.MS m/z(ESI):304.2[M+H]
3rd step, preparation 4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) aniline:
1-methyl-4-(4-nitro-2-(trifluoromethyl) benzyl) piperazine (5g) is dissolved in 65mL 75% ethanol, adds 0.5g 10% Palladium carbon, after nitrogen displacement, reactant liquor is stirred at room temperature 5h, after completion of the reaction filtration under diminished pressure under hydrogen, and filtrate obtains target after being evaporated Product, faint yellow solid.MS m/z(ESI):274.2[M+H].
4th step, preparation 3-iodo-4-methyl-N-(4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) phenyl) benzamide:
Intermediate 64, faint yellow solid (2.2g, yield is obtained according to method of condensing similar described in intermediate 33 the 5th step 74.3%).1H NMR(400MHz,DMSO-d6) δ 10.49 (s, 1H), 8.43 (s, 1H), 8.18 (d, J=1.5Hz, 1H), (8.04 d, J=8.5Hz, 1H), 7.92 (d, J=8.0Hz, 1H), 7.70 (d, J=8.5Hz, 1H), 7.50 (d, J=8.0Hz, 1H), 3.56 (s, 2H), 2.45 (s, 3H), 2.36 (d, J=20.7Hz, 8H), 2.16 (s, 3H).MS m/z(ESI):518.3[M+H].
The preparation of embodiment 65 intermediate 65-70
With the benzoic acid containing different substituents with aniline containing different substituents as raw material, use intermediate 64 or intermediate 36 institute State similar approach and obtain following intermediate 65-70.
Embodiment 713-acetenyl-4-methyl-N-(4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) phenyl) benzamide is (middle Body 71) preparation
The first step, the preparation iodo-methyl 4 methylbenzoate of 3-:
Iodo-for 3-4-ar-Toluic acid (15g, 57.14mmol) is dissolved with methanol, concentrated sulphuric acid is slowly dropped into wherein, reactant liquor Heat release, is warming up to 70 DEG C of reactions after dropping.TLC monitors, and after 48h, reaction is completely, stopped reaction, evaporated under reduced pressure methanol, Obtaining brown oil, be poured slowly into by grease in 200mL water, white milkiness shape, and heat release, water DCM extracts 2 times, DCM layer extracts once with sodium bicarbonate aqueous solution, saturated sodium-chloride water solution, water the most successively, and DCM layer is with anhydrous Sodium sulfate is dried, and obtains target product, yellowish-brown grease after evaporated under reduced pressure DCM.
Second step, preparation 3-trimethylsilyl acetylene base-methyl 4 methylbenzoate:
By upper step grease THF dissolve, add Hydro-Giene (Water Science). (1.1g, 5.74mmol, 0.1eq) and tetra-triphenylphosphine palladium (3.3g, 2.86mmol, 0.05eq), nitrogen is replaced three times, adds triethylamine (11.57g, 114.28mmol, 2.0eq) and trimethyl silicane Acetylene (8.4g, 85.71mmol, 1.5eq), stirring at normal temperature, react complete after 24h.Evaporated under reduced pressure solvent, column chromatography for separation Obtain target product, yellow oil.
3rd step, preparation 3-acetenyl-methyl 4 methylbenzoate:
Upper step grease methanol is dissolved, adds potassium carbonate (3.95g, 28.57mmol, 0.5eq), stirring at normal temperature 10min Rear reaction is complete, evaporated under reduced pressure solvent, and DCM dissolves, more successively with sodium bicarbonate aqueous solution, saturated sodium-chloride water solution, water Once, DCM layer anhydrous sodium sulfate is dried, and obtains target product, pale yellow oil through column chromatography for separation in extraction.
4th step, preparation 3-acetenyl-4-ar-Toluic acid:
The product methanol upper step obtained dissolves, and adds saturated sodium hydrate aqueous solution 10mL, and stirring at normal temperature is overnight.React Bi Hou, evaporated under reduced pressure solvent, add water about 20mL, with hydrochloric acid, pH furnishing is about 3, separates out a large amount of white solid, use DCM Extracting 3 times, merge DCM layer, wash DCM layer with water once, anhydrous sodium sulfate is dried, and evaporated under reduced pressure solvent is produced Product 3-acetenyl-4-ar-Toluic acid, faint yellow solid powder (8.0g, total recovery 87.2%).1H NMR(400MHz, DMSO-d6) δ 13.04 (br.s, 1H), 7.93 (s, 1H), 7.85 (d, J=8.0Hz, 1H), 7.43 (d, J=8.0Hz, 1H), 4.48 (s,1H),2.45(s,3H)。MS m/z(ESI):159.0[M-H]。
5th step, preparation 3-acetenyl-4-methyl-N-(4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) phenyl) benzamide:
3-acetenyl-4-ar-Toluic acid (500mg, 3.12mmol, 1.05eq) DCM is dissolved, addition HATU (1.42g, 3.74mmol, 1.2eq), DIPEA (806mg, 6.24mmol, 2.0eq), after stirring at normal temperature about 30min, add 3-trifluoro Methyl-4-[(4-methylpiperazine-1-yl) methyl] aniline (812mg, 2.97mmol, 1.0eq), is warming up to 45 DEG C of backflows, after 20h React complete.Evaporated under reduced pressure solvent, extracts 2 times with DCM and saturated sodium bicarbonate aqueous solution, and DCM layer obtains Huang after being evaporated Color grease, obtains intermediate 71, brown solid (807mg, yield about 65.4%) through column chromatography for separation.1H NMR(400 MHz,CDCl3) δ 8.33 (s, 1H), 7.91 (s, 1H), 7.86 (d, J=13.7Hz, 2H), 7.73 (t, J=7.1Hz, 2H), 7.28 (s, 1H),3.60(s,2H),3.32(s,1H),2.48(br.s,11H),2.30(s,3H)。MS m/z(ESI):416.3[M+H]。
The preparation of embodiment 72-76 intermediate 72-76
With the aniline containing different substituents or arylamine as raw material, use method similar described in intermediate 71 or intermediate 36 Obtain following intermediate 72-76.
Embodiment 773-acetenyl-4-methyl-N-(3-(4-methyl-1 H-imidazole-1-group)-5-(trifluoromethyl) phenyl) benzamide (intermediate 77) preparation
The first step, preparation 3-(4-methyl-imidazoles-1-base)-5-trifluoromethyl-aniline:
By bromo-for 3-5-trifluoromethyl-aniline (500mg, 2.1mmol, 1.0eq), 4-methyl isophthalic acid H-imidazoles (20.5mg, 2.5mmol, 1.2eq), Hydro-Giene (Water Science). (60mg, 0.3mmol, 0.14eq) and 8-hydroxyquinoline (44mg, 0.3mmol, 0.14eq) are molten Solution is in 3mL dimethyl sulfoxide, and nitrogen is warming up to 120 DEG C of reactions overnight after replacing 3 times, dilute after completion of the reaction, DCM Extracting, organic layer uses dilute ammonia solution successively, and saturated sodium-chloride water solution washs, and anhydrous magnesium sulfate is dried, and filters, reduces pressure dense After contracting, residue obtains target product, yellow solid (392mg, yield 77.3%) through column chromatography for separation.1HNMR(400MHz,CDCl3) δ7.73(s,1H),6.98(s,1H),6.92(s,1H),6.83(s,1H),6.77(s,1H),4.14(s,2H),2.27(s,3H)。MS m/z(ESI):242.1[M+H]。
Second step, preparation 3-acetenyl-4-methyl-N-(3-(4-methyl-1 H-imidazole-1-group)-5-(trifluoromethyl) phenyl) benzamide:
Method of condensing similar described in intermediate 33 the 5th step is used to obtain intermediate 77, faint yellow solid (350mg, yield 65.7%).1H NMR(400MHz,DMSO-d6)δ10.68(s,1H),8.29(s,1H),8.22(s,1H),8.16(s,1H), 8.12 (s, 1H), 7.93 (d, J=8.0Hz, 1H), 7.74 (s, 1H), 7.51 (d, J=9.0Hz, 2H), 4.56 (s, 1H), 2.48 (s, 3H),2.19(s,3H)。MS m/z(ESI):384.1[M+H]。
Embodiment 783-acetenyl-4-methyl-N-(3-((4-methylpiperazine-1-yl) methyl)-5-(trifluoromethyl) phenyl) benzamide is (middle Body 78) preparation
The first step, preparation 3-nitro-5-trifluoromethyl benzoyl chloride:
3-nitro-5-(Trifluoromethyl)benzoic acid. (3.52g, 14.97mmol, 1.0eq) is dissolved in 20mL thionyl chloride, and 80 DEG C are returned Stream 2h, after completion of the reaction evaporated under reduced pressure solvent, remaining residue is target product, is directly used in the next step.
Second step, preparation (4-methylpiperazine-1-yl) (3-nitro-5-(trifluoromethyl) phenyl) ketone:
N methyl piperazine (1.5g, 15mmol, 1.05eq), triethylamine (2.2g, 22.4mmol, 1.5mmol) is dissolved in 20ml DCM In, upper step product 3-nitro-5-trifluoromethyl benzoyl chloride is dissolved in 6mL DCM, is slowly added dropwise to reactant liquor, after finishing Room temperature reaction 0.5h, the most successively with saturated sodium bicarbonate solution, saturated nacl aqueous solution and water washing, anhydrous slufuric acid Magnesium is dried, and filters, is directly used in the next step after evaporated under reduced pressure.MS m/z(ESI):318.1[M+H]
3rd step, preparation 1-methyl-4-(3-nitro-5-(trifluoromethyl) benzyl) piperazine:
By (4-methylpiperazine-1-yl) (3-nitro-5-(trifluoromethyl) phenyl) ketone (14.97mmol) in anhydrous tetrahydro furan, Nitrogen is protected, and is cooled to 0 DEG C.With anhydrous tetrahydro furan by the borane dimethylsulf iotade (22.5mL, 45mmol, 3.0eq) of 2mol/L It is diluted to 50mL, is then slowly added dropwise to reactant liquor, be warming up to 65 DEG C of reactions after finishing overnight, be cooled to after completion of the reaction 0 DEG C, it is slowly added dropwise 6M HCl 22.5mL, is warming up to 65 DEG C of reaction 1h after finishing, is then cooled to 0 DEG C, is slowly added dropwise 4MNaOH aqueous solution adjusts pH to 9, and reactant liquor is extracted with ethyl acetate 3 times, merges organic layer, and anhydrous magnesium sulfate is dried, Filtration under diminished pressure, filtrate is evaporated rear residue and obtains target product, pale yellow oil (2.4g, yield 53.2%) through column chromatography for separation. MS m/z(ESI):304.1[M+H]
4th step, preparation 3-((4-methylpiperazine-1-yl) methyl)-5-(trifluoromethyl) aniline:
1-methyl-4-(3-nitro-5-(trifluoromethyl) benzyl) piperazine (2.4g, 7.96mmol) is dissolved in 75% ethanol, adds 10% Palladium carbon (240mg, 10%eq), after nitrogen displacement, logical hydrogen reacts at 50 DEG C, and after completion of the reaction, filtration under diminished pressure, filtrate is steamed Do to obtain target product, white solid (2.1g, yield 96.1%).MS m/z(ESI):274.1[M+H]
5th step, preparation 3-acetenyl-4-methyl-N-(3-((4-methylpiperazine-1-yl) methyl)-5-(trifluoromethyl) phenyl) benzamide:
Method of condensing similar described in intermediate 33 the 5th step is used to obtain intermediate 78, faint yellow solid (260mg, hydrochlorate, Yield 61.7%).1H NMR(400MHz,DMSO-d6)δ10.55(s,1H),9.28(br.s,1H),8.10(s,3H),7.91(d, J=8.1Hz, 1H), 7.49 (d, J=8.1Hz, 1H), 7.41 (s, 1H), 4.56 (s, 1H), 3.66 (s, 2H), 3.39 (d, J=11.1 Hz, 2H), 3.14-2.99 (m, 2H), 2.94 (d, J=12.5Hz, 2H), 2.80 (s, 3H), 2.47 (s, 3H), 2.33 (t, J=11.9 Hz,2H)。MS m/z(ESI):416.3[M+H]。
The preparation of embodiment 793-acetenyl-N-(3-(trifluoromethyl) phenyl) benzamide (intermediate 79)
The first step, the preparation iodo-N-of 3-(3-(trifluoromethyl) phenyl) benzamide:
By m-iodobenzoic acid (1.29g, 5.2mmol, 1.05eq), TBTU (O-(benzotriazole-1-oxygen)-N, N, N ', N '-four MU hexafluoro borate) (1.93g, 6.0mmol, 1.2eq), triethylamine (1.01g, 10mmol, 2.0eq) addition DCM In, stirring at normal temperature 0.5h, add 3-Aminotrifluorotoluene (805mg, 5.0mmol, 1.0eq), be warming up to 45 DEG C of backflows.12h Rear reaction is complete.Evaporated under reduced pressure solvent, extracts once with saturated sodium bicarbonate aqueous solution, DCM, and DCM layer is evaporated, through post Chromatography obtains target product.1H NMR(400MHz,DMSO-d6)δ10.60(s,1H),8.33(s,1H),8.22(s, 1H), 8.05 (d, J=8.0Hz, 1H), 7.99 (dd, J=7.8,1.2Hz, 2H), 7.61 (t, J=8.0Hz, 1H), 7.47 (d, J=7.8 Hz, 1H), 7.37 (t, J=8.0Hz, 1H).MS m/z(ESI):392.0[M+H].
Second step, preparation N-(3-(trifluoromethyl) phenyl)-3-((trimethyl silyl) acetenyl) benzamide:
Upper step product is dissolved in THF (oxolane), adds Hydro-Giene (Water Science). (95.23mg, 0.5mmol, 0.1eq), double triphen Base phosphine dichloride palladium (175.5mg, 0.25mmol, 0.05eq), nitrogen replace three times, add DIPEA (1.29g, 10mmol, 2.0eq), trimethylsilyl acetylene (737mg, 7.5mmol, 1.5eq), stirring at normal temperature, react complete after 4h.Stopped reaction, Obtain target product through column chromatography for separation, be directly used in the next step.MS m/z(ESI):362.2[M+H].
3rd step, preparation 3-acetenyl-N-(3-(trifluoromethyl) phenyl) benzamide:
Upper step product with methylalcohol is dissolved, adds potassium carbonate (345.5mg, 2.5mmol, 0.5eq), stirring at normal temperature, 10min Rear reaction is complete.Solvent evaporated, residue obtains intermediate 79, faint yellow solid (875.0mg, yield through column chromatography for separation About 60.5%).1H NMR(400MHz,DMSO-d6) δ 10.61 (s, 1H), 8.25 (s, 1H), 8.10 (s, 1H), 8.06 (d, J= 8.3Hz, 1H), 8.00 (d, J=7.9Hz, 1H), 7.72 (d, J=7.7Hz, 1H), 7.63 7.56 (m, 2H), 7.47 (d, J=7.9Hz, 1H),4.35(s,1H).MS m/z (ESI): 290.1 [M+H].
The preparation of embodiment 80 intermediate 80
With the halogen benzoic acid containing different substituents with the aniline containing different substituents as raw material, use described in intermediate 79 similar Method obtains following intermediate 80.1H NMR(400MHz,CDCl3) δ 7.98 (s, 1H), 7.91 (s, 1H), 7.85 (d, J= 7.6Hz, 1H), 7.77 (d, J=7.2Hz, 1H), 7.46 (d, J=8.4Hz, 2H), 7.44-7.39 (m, 2H), 3.43 (s, 1H).MS M/z (ESI): 324.1 [M+H].
The preparation of embodiment 811-(3-ethynyl phenyl)-3-(3-(trifluoromethyl) phenyl) urea (intermediate 81)
Triphosgene (1.79g, 6.0mmol, 1.5eq) is dissolved in EA, stirring at normal temperature.By 3-Aminotrifluorotoluene (967mg, 6.0mmol, 1.5eq) it is dissolved in EA, is placed in Dropping funnel, be slowly dropped in the EA solution of triphosgene, drip complete after 30min.So After be slowly added to triethylamine (1.2g, 12mmol, 2.0eq), separate out a large amount of white solid, normal-temperature reaction.After 2h, decompression is steamed Dry solvent, adds EA, filters off insoluble matter, collects filtrate.In filtrate add between acetylenylaniline (469mg, 4.0mmol, 1.0eq), there is solid to produce.TLC monitors after completion of the reaction, and volume ratio ether used by evaporated under reduced pressure solvent: oil Ether=1:1 is recrystallized to give intermediate 81, white solid (543mg, yield about 44.6%).1H NMR(400MHz,DMSO-d6) δ 9.35 (s, 1H), 9.15 (s, 1H), 8.01 (s, 1H), 7.68 (s, 1H), 7.59 (d, J=8.4Hz, 1H), 7.52 (t, J=7.9Hz, 1H), 7.44 (d, J=8.2Hz, 1H), 7.31 (d, J=7.8Hz, 2H), 7.10 (d, J=7.6Hz, 1H), 4.17 (s, 1H).MS m/z(ESI):304.2[M+H]。
The preparation of embodiment 82-83 intermediate 82-83
With the aniline containing different substituents as raw material, method similar described in intermediate 81 is used to obtain following intermediate 82-83.
Embodiment 84N-(3-((4-amino-1-isopropyl-1H-pyrazolo [3,4-d] pyrimidin-3-yl) acetenyl)-4-aminomethyl phenyl)-4-((4- Methylpiperazine-1-yl) methyl) preparation of benzamide (compound 1)
N-(3-iodo-4-aminomethyl phenyl)-4-((4-methylpiperazine-1-yl) methyl) benzamide (118mg, 0.263mmol, 1.0eq), 3-second Alkynyl-1-isopropyl-1H-pyrazolo [3,4-d] pyrimidine-4-amine (55.57mg, 0.276mmol, 1.05eq), CuI (5.7mg, 0.03mmol, 0.1eq), PdCl2(PPh3)2(21.06mg, 0.03mmol, 0.1eq), in there-necked flask, adds 2mL DMF molten After solution, nitrogen is replaced three times, adds triethylamine (53.23mg, 0.526mmol, 2.0eq) in 80 DEG C of reactions overnight, and TLC monitors After completion of the reaction, decompression boils off DMF, and residue obtains compound 1, faint yellow solid (62mg, yield through column chromatography for separation 45.1%).1H NMR(400MHz,DMSO-d6) δ 10.33 (s, 1H), 8.27 (s, 1H), 8.10 (s, 1H), 7.94 (d, J= 7.6Hz, 2H), 7.75 (d, J=8.2Hz, 1H), 7.47 (d, J=7.6Hz, 2H), 7.34 (d, J=8.2Hz, 1H), 5.20-4.93 (m, 1H), 3.56 (s, 2H), 2.58 (br.s, 8H), 2.47 (s, 3H), 2.34 (s, 3H), 1.49 (d, J=6.4Hz, 6H) .MS m/z(ESI):523.2930[M+H]。
The preparation of embodiment 85-175 compound 2-92
With the aryl ethane containing different substituents and halides as raw material, method similar described in embodiment 84 is used to obtain compound 2-92。
Embodiment 933-((4-amino-1-(piperidin-4-yl)-1H-pyrazolo [3,4-d] pyrimidin-3-yl) acetenyl)-4-methyl-N-(4-((4-first Base piperazine-1-base) methyl)-3-(trifluoromethyl) phenyl) preparation of benzamide (compound 93)
Tert-butyl-4-(4-amino-3-((2-methyl-5-((4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) phenyl) carbamyl) Phenyl) acetenyl)-1H-pyrazolo [3,4-d] pyrimidine-1-base) piperidines-1-carboxylate (1.46g, 2mmol) is dissolved in 20mL DCM, Add 10ml trifluoroacetic acid after being cooled to 0 DEG C, be naturally warmed to room temperature reaction, after 0.5h, react complete, evaporated under reduced pressure solvent, residual Adjust PH to 8 after excess water dissolution, separate out a large amount of solid, filtration under diminished pressure, wash filter cake, after solid is vacuum dried, obtain chemical combination Thing 93, pale yellow powder (1.2g, yield 94.9%).
1H NMR(400MHz,DMSO-d6)δ10.56(s,1H),8.35(s,1H),8.28(s,1H),8.22(s,1H),8.08 (d, J=7.9Hz, 1H), 7.98 (d, J=7.9Hz, 1H), 7.72 (d, J=8.2Hz, 1H), 7.55 (d, J=8.2Hz, 1H), 5.09-4.98 (m, 1H), 4.36-4.32 (m, 1H), 3.58 (s, 2H), 3.21-3.08 (m, 2H), 2.99 (d, J=7.0Hz, 2H), 2.58 (s, 3H), 2.42 (br.s, 8H), 2.31 (d, J=11.7Hz, 2H), 2.22 (s, 3H), 2.12 (d, J=11.6Hz, 2H).MS m/z(ESI):632.3071[M+H]。
Embodiment 943-((1-(1-Antiepilepsirin-4-base)-4-amino-1H-pyrazolo [3,4-d] pyrimidin-3-yl) acetenyl)-4-methyl The preparation of-N-(4-((4-methylpiperazine-1-yl) methyl)-3-(trifluoromethyl) phenyl) benzamide (compound 94)
3-((4-amino-1-(piperidin-4-yl)-1H-pyrazolo [3,4-d] pyrimidin-3-yl) acetenyl)-4-methyl-N-(4-((4-methyl piperazine -1-base) methyl)-3-(trifluoromethyl) phenyl) benzamide (510mg, 0.807mmol, 1.0eq) is dissolved in 10mL DCM, adds Enter triethylamine (163mg, 1.6mmol, 2.0eq), be cooled to 0 DEG C, be slowly added dropwise acryloyl chloride (72 μ L, 0.888mmol, Dichloromethane solution (4mL) 1.1eq), after finishing, reaction is completely.The most successively with saturated aqueous ammonium chloride, unsaturated carbonate hydrogen Sodium water solution, saturated sodium-chloride water solution washing reaction liquid, remaining organic layer anhydrous magnesium sulfate is dried, and filters, filtrate decompression Being evaporated, residue is through acetone: ether (1:1) washs to obtain compound 94, pale yellow powder (440.1mg, yield 72.3%).
1H NMR(400MHz,DMSO-d6)δ10.54(s,1H),8.33(s,1H),8.27(s,1H),8.21(s,1H),8.06 (d, J=7.7Hz, 1H), 7.96 (d, J=7.7Hz, 1H), 7.71 (d, J=7.8Hz, 1H), 7.54 (d, J=7.8Hz, 1H), 6.88 (dd, J=17.7,10.2Hz, 1H), 6.14 (d, J=17.7Hz, 1H), 5.71 (d, J=10.2Hz, 1H), 5.08-4.94 (m, 1H), 4.56 (d, J=10.5Hz, 1H), 4.21 (d, J=10.9Hz, 1H), 3.57 (s, 2H), 2.97-2.84 (m, 1H), 2.73-2.62 (m, 1H), 2.57 (s, 3H),2.39(br.s,8H),2.19(s,3H),2.01(br.s,4H)。MS m/z(ESI):686.3181[M+H]。
Embodiment 95 kinase inhibition assay
The purpose of this experiment is the detection the compounds of this invention inhibitory activity to vitro kinase, and the method for employing is isotope-labelling method (the γ phosphate group on labelling ATP).This experiment respectively to Abl (T315I) (h), ALK (h), ARK5 (h), Axl (h), Blk (h), Bmx(h)、BTK(h)、B-Raf(h)、cKit(h)、cSRC(h)、CDK7、CHK1(h)、c-RAF(h)、DDR2(h)、EGFR(h)、 EphA1(h)、EphA2(h)、EphA8(h)、EphB2(h)、EphB4(h)、ErbB2(h)、FAK(h)、Fer(h)、FGFR1(h)、 Flt3(h)、Fms(h)、Fyn(h)、Hck(h)、GSK3β(h)、IKKα(h)、IKKβ(h)、Itk(h)、JAK3(h)、JNK1 α1(h)、KDR(h)、Lyn(h)、MAPK1(h)、MEK1(h)、Met(h)、mTOR(h)、PAK1(h)、PDGFRα(h)、 Pim-1(h)、PKA(h)、PKBα(h)、PKBβ(h)、PKCα(h)、Ret(h)、RIPK2(h)、Src(1-530)(h)、TAK1(h)、 The kinases such as TBK1 (h), Tec (h) activated, Tie2 (h), TrkA (h), ULK3 (h), Yes (h), PI3Kinase a (h) are carried out External activity suppression test.The kinase inhibiting activity IC of test-compound50(half-inhibition concentration) or test-compound are 10 Under μM concentration, the suppression ratio to kinase activity represents.IC50Value can pass through test-compound under a series of variable concentrations to swashing The suppression ratio of enzymatic activity calculates and obtains.Experimental technique is as follows: in a reaction tube, be sequentially added into buffer (8mM MOPS, pH 7.0,0.2mM EDTA,10mM MnCl2), kinases to be measured (5-10mU), kinase whose substrate to be measured, the acetic acid of 10mM Magnesium and γ 33P-ATP solution, and the test-compound of variable concentrations.Then in reaction, MgATP is added to start enzyme reaction Process, and at room temperature hatch 40 minutes.Finally terminate reaction with 3% phosphate buffer of 5 μ l, and the reaction of 10 μ L Liquid is titrated on Filtermat A film, washes three times with the phosphate solution of 75mM, and each 5 minutes, again with methanol was washed once. Finally being dried Filtermat A film and it is carried out scinticounting, the size of scintillascope numerical value reflects the degree that substrate is phosphorylated, Such that it is able to characterize kinase activity to be suppressed situation.
Table 1 gives the part test-compound IC50 value to part kinase inhibiting activity.(all tables of "--" in following table Show and test.)
Table 1 part test-compound is to various kinase whose inhibitory activity (IC50:nM)
Kinases Compound 3 Compound 31 Compound 93 Compound 94
Abl <1 -- -- --
Abl(T315I) -- -- -- 133
c-Src(1-530) <1 2 -- --
c-Src(T341M) 30 3733 -- --
B-Raf(V600E) 15 5 -- --
B-Raf 92 22 -- --
c-RAF 27 19 -- --
Yes <1 <1 <1 2
Fyn 5 3 -- --
Blk 19 12 -- --
Bcr-Abl <1 -- -- --
KDR 17 12 -- --
FGFR1 3 -- -- --
EphA2 16 18 13 --
EphB2 26 10 -- --
EphB4 23 11 28 --
ErbB2 -- 251 -- --
DDR1 9 4 -- --
DDR2 128 74 -- --
TAK1 61 203 -- --
TrkA 27 -- 91 52
Btk 67 127 -- --
Bmx 6 18 11 22
IKKα 353 -- -- --
IKKβ 164 -- -- --
Axl 578 -- -- --
PDGFRα 890 396 -- --
JAK2 2911 -- -- --
EGFR 3518 -- -- --
Result shows, part test-compound to kinases such as Abl, Abl (T315I), c-Src (1-530), c-Src (T341M), B-Raf(V600E)、B-Raf、c-RAF、Yes、Fyn、Blk、Bcr-Abl、KDR、FGFR1、EphA2、EphB2、 EphB4, ErbB2 (h), DDR1, DDR2, TAK1, TrkA, Btk, Bmx have preferable inhibitory activity, swash part Enzyme such as IKK α, IKK β, Axl, PDGFR α have medium inhibitory activity.
Table 2 gives the part test-compound IC to c-Src (1-530) kinase inhibiting activity50Value.
Table 2 part test-compound inhibitory activity kinase whose to c-Src (IC50:nM)
Compound IC50(nM) Compound IC50(nM)
1 4 17 4
5 2 18 4
6 9 19 8
7 19 20 7
8 7 21 7
9 15 24 59
10 83 25 3191
11 10 26 11
12 139 29 5
13 11 30 20
14 3 93 12
16 7 94 15
Result shows, part test-compound has preferable inhibitory activity to Src kinases.
Table 3 give part test-compound under the concentration of 10 μMs respectively to Abl (T315I) (h), ALK (h), ARK5 (h), Axl(h)、Blk(h)、Bmx(h)、BTK(h)、B-Raf(h)、cKit(h)、cSRC(h)、CDK7、CHK1(h)、c-RAF(h)、 DDR2(h)、EGFR(h)、EphA1(h)、EphA2(h)、EphA8(h)、EphB2(h)、EphB4(h)、ErbB2(h)、FAK(h)、 Fer(h)、FGFR1(h)、Flt3(h)、Fms(h)、Fyn(h)、Hck(h)、GSK3β(h)、IKKα(h)、IKKβ(h)、Itk(h)、 JAK3(h)、JNK1α1(h)、KDR(h)、Lyn(h)、MAPK1(h)、MEK1(h)、Met(h)、mTOR(h)、PAK1(h)、 PDGFRα(h)、Pim-1(h)、PKA(h)、PKBα(h)、PKBβ(h)、PKCα(h)、Ret(h)、RIPK2(h)、 Src(1-530)(h)、TAK1(h)、TBK1(h)、Tec(h)activated、Tie2(h)、TrkA(h)、ULK3(h)、Yes(h)、 The suppression ratio (numeric representation relative activity percent protein) of PI3Kinase a (h) kinase activity.
Table 3 part test-compound is suppression ratio kinase whose to part under the concentration of 10 μMs
Result shows, part test-compound to Abl (T315I), Axl, Blk, Bmx, BTK, B-Raf, cSRC, c-RAF, DDR2、EphA1、EphA2、EphA8、EphB2、EphB4、ErbB2、FGFR1、Flt3、Fms、Fyn、Hck、 IKKα、IKKβ、Itk、JAK3、KDR、Lyn、PDGFRα、Ret、RIPK2、Src(1-530)、TAK1、Tec、 Tie2, TrkA, ULK3, Yes have preferable inhibitory activity.Part test-compound to ALK, cKit, EGFR, FAK, Fer, JNK1 α 1, PKA, PKB β have medium inhibitory activity.
Embodiment 96 cell growth inhibition assay
The purpose of this experiment be detection the compounds of this invention to vitro human tumor cell proliferation inhibition activity, the method for employing is MTT (tetramethyl azo azoles salt) colorimetry.
1) experiment material:
Main agents: RPMI-1640, hyclone, pancreatin etc. purchased from Gibco BRL company (Invitrogen Corporation, USA), DMEM culture medium is purchased from ATCC (American Type Culture Collection).Tetramethyl azo azoles salt (MTT), Dimethyl sulfoxide (DMSO) is Sigma company (USA) product.Compound 3 and compound 31 are synthesized by inventor, body It is configured to 10mM with 100%DMSO during outer experiment and stores liquid, put-20 DEG C of refrigerators and keep in Dark Place standby, face the used time with training completely Nutrient solution is diluted to desired concn.
Cell line and cultivation: Breast cancer lines MDA-MB-231 used by this experiment, MCF-7, SKBR-3, BT474, MDA-MB-468, MDA-MB-453, MDA-MB-435, large B cell lymphoid tumor cell strain LY-10, HBL-1, people Pancreas cancer cell strain Panc-1, Miapaca-2, human lung carcinoma cell line A549, H358, human leukemia cell line THP-1, people Hepatoma cell strain Hepg2, human melanoma cell strain A2058 etc. are all purchased from U.S. ATCC (American type culture Collection), this laboratory preserve.All above human lymphoma cell's strain, large B cell lymphoid tumor cell strain, T cell are drenched Bar tumor cell strain is trained completely with containing 10% hyclone, 100U/mL penicillin, the RPMI-1640 of 100 μ g/mL streptomycins Support base at 5%CO2, cultivate under the conditions of 37 DEG C.Remaining cell strain use containing 10% hyclone (MV4-11 cell is 20%), 100U/ml penicillin, 100 μ g/mL streptomycins DMEM complete medium at 5%CO2, cultivate under the conditions of 37 DEG C.
2) experimental technique:
Adjusting cell concentration with complete cell culture fluid is 1～2 × 104The cell suspension of individual/mL, is inoculated in 96 orifice plates, every hole 200 μ L cell suspension, overnight incubation.Next day, inhale and abandon supernatant (suspension cell draws supernatant after being centrifuged), use gradient the most respectively The test-compound of concentration processes cell.Set negative control group and isopyknic solvent control group, the DMSO of not drug containing simultaneously Concentration is 0.1%, and each dosage group sets 3 multiple holes, at 37 DEG C, 5%CO2Under the conditions of cultivate.After 72 hours, every hole adds Concentration is the MTT reagent 20 μ L of 5mg/mL, after being further cultured for 2-4h, abandons supernatant, and every hole adds DMSO 150 μ L, Vibration mixing 15min, measures absorbance (A) value (A value is directly proportional) to viable count by microplate reader (λ=570nm), Take its meansigma methods.Relative cell proliferation suppression ratio=(control group A 570-experimental group A570) × 100%/control group A 570.Real Test and be at least repeated 3 times.Experimental data mean represents, data statistics data uses t inspection, and P < 0.05 has statistics for difference Meaning.Following compound on intracellular inhibited proliferation all uses IC50Or suppression ratio represents.
3) experimental result:
Use above method, to Breast cancer lines MDA-MB-231, MCF-7, SKBR-3, BT474, MDA-MB-468, MDA-MB-453, MDA-MB-435, large B cell lymphoid tumor cell strain LY-10, HBL-1, people Pancreas cancer cell strain Panc-1, Miapaca-2, human lung carcinoma cell line A549, H358, human leukemia cell line THP-1, people Hepatoma cell strain Hepg2, human melanoma cell strain A2058 etc. have carried out proliferation inhibition activity test, the results are shown in Table 4.
The table 4 part test-compound proliferation inhibition activity (IC to various cell strains50:μM)
Cell strain Compound 3 Compound 31 Cell strain Compound 3 Compound 31
HBL-1 ～10 -- Hela 5.41 7.23
OCI-LY10 2.168 -- PC-3 -- 3.974
RAMOS ～10 -- MDA-MB-231 0.0113 0.01533
MV4-11 0.020 -- MDA-MB-435 0.008654 0.0147
THP-1 5.11 7.378 MDA-MB-453 3.369 --
PANC-1 6.556 -- MDA-MB-468 4.478 3.216
Miapaca-2 0.01277 0.04036 BT474 >10 >10
CFPAC 0.607 0.4721 MCF-7 -- 1.987
H4 -- 1.807 SKBR-3 4.523 1.265
U87 0.742 5.420 MM.1S 0.872 2.383
A2058 1.908 -- HCT116 0.178 0.2303
H358 1.665 -- HT29 0.1663 0.3496
A549 0.1988 1.423 SW1990 0.3559 0.2061
HepG2 8.672 6.167 A375 0.02088 0.02657
plc/prf/5 -- 3.974
Result shows, test-compound 3 and compound 31 to MV4-11, Miapaca-2, MDA-MB-231, MDA-MB-435, A375 cell has stronger inhibitory activity；Test-compound 3 and compound 31 are to other tumor cell Strain includes that CFPAC, U87, MM.1S, HCT116, HT29, A549 etc. also have medium inhibitory activity.
Part test-compound is shown in Table 5 to the proliferation inhibition activity of MDA-MB-231 and MDA-MB-435 cell.Wherein, IC50< 100nM symbol ++++represent, 100nM < IC50< 500nM symbol +++ represent, 500nM < IC50< 1000nM uses Symbol ++ represent, IC50> 1000nM symbol+represent.
Table 5 compound proliferation inhibition activity to MDA-MB-231 and MDA-MB-435 cell
Result shows, the proliferation inhibition activity that part test-compound is stronger to MDA-MB-231 and MDA-MB-435 cell.
Embodiment 97 compound 3 and the compound 31 internal pharmacodynamic experiment to SCID nude mice
The purpose of this experiment is the anti-tumor in vivo effect of detection the compounds of this invention.This experiment uses SCID mice Subcutaneous tumor Model, the anti-tumor in vivo activity of test invention compound 3 and compound 31.Cell strain used is breast carcinoma cell strain MDA-MB-231.With the anti-breast cancer medicines paclitaxel being currently in use and breast carcinoma clinic at the marketed drug Dasatinib ground it is Positive control.
1) experiment material:
Hyclone, pancreatin etc. are purchased from Gibco BRL company (Invitrogen Corporation, USA), DMEM culture medium Purchased from ATCC (American Type Culture Collection), Breast cancer lines MDA-MB-231 is purchased from U.S. ATCC Company, SCID mice is purchased from Fukang biotech inc of China of BeiJing, China.Paclitaxel is vast fragrant biological purchased from Chinese Shanghai Technology Co., Ltd..Dasatinib is purchased from Nanjing of China Kang Manlin chemical industry Industrial Co., Ltd..
2) experimental technique:
Use 6～8 week old SCID mice, according to about 5 × 106Individual/0.1mL/ MDA-MB-231 cell concentration is inoculated in little Flank after under Corium Mus, treats that tumor grows to 200mm3Afterwards (about 15 days), mice group (n=6) and start be administered.Respectively organize medicine It is dissolved in 5%DMSO+25%PEG-400+70% water.
Experiment packet one:
Drug solvent matched group, every day oral administration gavage blank solvent 200uL；
Compound 3 according to dosage 30mg/kg oral administration gavage every day is administered；
Compound 3 according to dosage 15mg/kg oral administration gavage every day is administered.；
Compound 3 according to dosage 7.5mg/kg oral administration gavage every day is administered；
Positive control Taxol according to dosage 10mg/kg tail vein injection weekly is administered；
Experiment packet two:
Drug solvent matched group, every day, oral administration gavage was administered blank solvent 200uL；
Compound 31 according to dosage 40mg/kg oral administration gavage every day is administered；
Compound 31 according to dosage 20mg/kg oral administration gavage every day is administered；
Compound 31 according to dosage 10mg/kg oral administration gavage every day is administered；
Positive control Taxol according to dosage 10mg/kg tail vein injection weekly is administered；
Positive control Dasatinib according to dosage 40mg/kg oral administration gavage every day is administered.
Observation index: measure Mouse Weights and tumor major diameter, minor axis and calculate gross tumor volume (length × width for every three days2 ×0.5).Observe each group of mice every day with or without reactions such as diarrhoea, tic, erythra, body weight substantially reductions.
3) experimental result:
The tumor growth curve of the experiment packet one that experiment records is shown in the tumor growth curve testing packet two that Fig. 1, experiment record See figure Fig. 2.
Test result indicate that, test-compound 3 has the suppression of obvious tumor growth and makees breast carcinoma cell strain MDA-MB-231 With, under the dosage of 30mg/kg every day, can substantially suppress tumor growth, and show and be better than positive control (paclitaxel) Inhibition.Compound 31 has obvious tumor growth inhibitory action to breast carcinoma cell strain MDA-MB-231, often Under the dosage of it 20mg/kg, can substantially suppress tumor growth, and show and be better than positive control (paclitaxel and Dasatinib) Inhibition.Administration process not finding, the untoward reaction such as body weight reduction, erythra, diarrhoea occurs in mice, shows at test agent Under amount, test-compound 3 and compound 31 toxicity in the range of dosage are relatively low.
The inhibitory activity test of the transgenic zebrafish angiogenesis of embodiment 98 compound 31
The purpose of this experiment is the detection invention compound inhibitory activity to internal new vessels, and the method for employing is that investigation is multiple dense The lower the compounds of this invention suppression situation to fluorescent transgenic Brachydanio rerio FLK1-GFP body intersegmental blood vessel of degree.Test-compound new Angiogenic inhibitory activity test-compound under 10ug/mL, 5ug/mL, 2.5ug/mL concentration to Brachydanio rerio body intersegmental blood vessel Suppression degree represents.With breast carcinoma clinical at the marketed drug Dasatinib ground as positive control.
1) experiment material:
Transgenic zebrafish (FLK1-GFP): this Laboratory culture
Experiment reagent: dimethyl sulfoxide (DMSO)；Test-compound；Dasatinib
Major experimental instrument: fluorescence microscope；Stereomicroscope；CCD camera etc..
2) experimental technique:
The acquisition of zebrafish embryo: the Brachydanio rerio used by laboratory is blood vessel fluorescent transgenic Brachydanio rerio (FLK1:GFP).Speckle The cultivation of horse fish and the method for cultivation reference Westerfield.In the previous day taking ovum, male and female Brachydanio rerio is pressed 1:1 proportional paired. Within second day, its natural mating is made to lay eggs at the temperature of about 28 DEG C and the illumination of abundance.Gather enough zebrafish embryos, clearly It is put in embryo medium after washing, and puts into 28 DEG C of incubators cultivations.Form and auxology standard is utilized to identify survival at any time Situation, embryo in heaven is white, should take out in time to prevent water qualitative change bad.
Drug treating: after zebrafish embryo after fertilization 10h (basis of microscopic observation Zebrafish Embryo is to the bud phase), with Embryo's packet of health chosen by machine, adds 24 orifice plates, 10, every hole zebrafish embryo, and exhaust culture fluid, adds different dense Degree compound solution.Compound 31 concentration arranges and is respectively 10ug/mL, 5ug/mL, 2.5ug/mL.Dasatinib concentration sets Put respectively 10ug/mL.Set blank simultaneously, be added without any compound.Then put it into 28 DEG C of incubators to cultivate. Result is observed: after zebrafish embryo fertilization 31h, takes out Brachydanio rerio and shells ovum.Then it is put on microscope slide, is added 1 ‰ Tricaine solution anesthesia fish body also fixes fish body, then to body intersegmental blood vessel (Inter under fluorescence microscope with 1.5% methylcellulose Segmental vessel, ISV) carry out observe counting take pictures.
3) experimental result:
Fig. 3 reflection is the compound 31 vascular study situation to FLK1 transgenic zebrafish under variable concentrations.Result shows, Comparing with matched group, compound 31 can suppress the angiogenesis of Brachydanio rerio well.Originally test result indicate that, the embodiment of the present invention In the compound 31 good inhibitory activity of new vessels to FLK1 transgenic zebrafish for preparing, this result reflects chemical combination Thing 31 has good inhibitory activity to VEGFR2.

Claims (16)

1.3-acetenyl Pyrazolopyrimidine derivative, its structure is as shown in formula I:
Wherein, R1For-H, C1～C4Alkyl,
R2For-H, C1～C8Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,
R3～R7Independently be-H, C1～C8Alkyl ,-OH, C1～C8Alkoxyl, halogen,
R8～R11Independently be-H, C1～C8Alkyl, halogen ,-OH,
R12～R14Independently be
R15～R19Independently be-H, C1～C8Alkyl ,-OH, C1～C8Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, halogen, C1～C8Alkyl, C1～C8Cycloalkyl ,-OCF3Or-CF3；
R39～R42Independently be C1～C8Alkyl, C3～C8Cycloalkyl or C1～C8Hydroxy alkyl；
N=0～6.
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 1, it is characterised in that:
R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,
R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen,
R8～R11Independently be-H, C1～C4Alkyl, halogen ,-OH,
R12～R14Independently be
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, halogen, C1～C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3；
R39～R42Independently be C1～C4Alkyl, C3～C8Cycloalkyl or C1～C4Hydroxy alkyl；
N=0～4；
Preferably, R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,
R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen,
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R12～R14Independently be
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, halogen, C1～C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3；
R39～R42Independently be C1～C4Alkyl, C3～C8Cycloalkyl or C1～C4Hydroxy alkyl；
N=0～4；
It is further preferred that R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,
R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen,
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R12～R14Independently be
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, halogen, C1～C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3；
R39～R42Independently be C1～C4Alkyl, C3～C8Cycloalkyl or C1～C4Hydroxy alkyl；
N=0～4；
Further preferred, R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,
R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen,
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R12～R14Independently be
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, halogen, C1～C4Alkyl, C1～C4Cycloalkyl ,-OCF3Or-CF3；
N=0～2；
Optimum, R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,
R3～R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl ,-F ,-Cl,
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R12～R14Independently be
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl ,-F ,-Cl ,-CF3、-OCF3、
R20～R38Independently be-H, C1～C4Alkyl or-CF3；N=0 or 1.
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 2, it is characterised in that: work as R6ForTime, Its structure is as shown in formula II:
Wherein, R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,N=0～4；
R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R12For
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, C1～C4Alkyl or-CF3。
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 3, it is characterised in that: R1For-H or
Preferably, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,N=0～4；R8～R11Independently be -H、C1～C4Alkyl ,-OH,
It is further preferred that R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl, N=0～4；R8～R11Independently be-H, C1～C4Alkyl ,-OH,
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl, N=0～2；R8～R11Independently be-H, C1～C4Alkyl ,-OH,
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl, N=0 or 1；R8～R11Independently be-H, C1～C4Alkyl ,-OH,
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl or halogen；
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl ,-F or-Cl；
Further preferred, R3～R5、R7Independently be-H, methyl or-Cl；
Preferably, R15～R19Independently be-H, C1～C4Alkyl, C1～C4Alkoxyl, halogen ,-CF3、-OCF3Or
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, methoxyl group ,-F ,-Cl ,-CF3、-OCF3Or
Optimum, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl, N=0 or 1；R3～R5、R7Independently be-H, methyl Or-Cl；R8～R11Independently be-H, C1～C4Alkyl ,-OH, R15～R19Independently be-H, C1～C4Alkyl, Methoxyl group ,-F ,-Cl ,-CF3、-OCF3OrR20～R38Independently be-H, C1～C4Alkyl or-CF3。
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 2, it is characterised in that: work as R6ForTime, Its structure is as shown in formula III:
Wherein, R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,N=0～4；
R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R13For
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, C1～C4Alkyl or-CF3。
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 5, it is characterised in that:
R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8、R9Independently be-H, C1～C4Alkyl ,-OH,
It is further preferred that R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3~ C8Epoxy alkyl,R8、R9Independently be-H, C1～C4Alkane Base ,-OH,
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl ,-OH,
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl,
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or halogen；
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or-Cl；
Preferably, R13For
Preferably, R15～R19Independently be-H, C1～C4Alkyl, C1～C4Alkoxyl, halogen ,-CF3Or
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, halogen ,-CF3Or
Further preferred, R15～R19Independently be-H, C1～C4Alkyl ,-CF3Or
Preferably, R20～R38Independently be-H or C1～C4Alkyl；
Optimum, R1For-H orC1～C4Alkyl,R2For C1～C4Alkyl, R8Substituted C3~ C8Cycloalkyl,C3～C8Epoxy alkyl,R3～R5、R7Independently be-H, C1~ C4Alkyl ,-OH or-Cl；R8、R9Independently be-H, C1～C4Alkyl,R13For R15～R19Independently be-H, C1～C4Alkyl ,-CF3OrR20～R38Independent Ground is-H or C1～C4Alkyl.
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 5, it is characterised in that: work as R13For Time, its structure is as shown in formula IV:
Wherein, R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,N=0～4；
R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 7, it is characterised in that:
R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl, N=0～4；R8、R9Independently be-H, C1～C4Alkyl ,-OH,
It is further preferred that R2For C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3~ C8Epoxy alkyl,R8、R9Independently be-H, C1～C4Alkane Base ,-OH,
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl ,-OH,
Further preferred, R2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8Alkylene oxide Base,R8、R9Independently be-H, C1～C4Alkyl,
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or halogen；
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or-Cl；
Preferably, R15～R19Independently be-H, C1～C4Alkyl, C1～C4Alkoxyl, halogen ,-CF3Or
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, halogen ,-CF3Or
Further preferred, R15～R19Independently be-H, C1～C4Alkyl ,-CF3Or
Optimum, R1For-H orR2For C1～C4Alkyl, R8Substituted C3～C8Cycloalkyl,C3～C8 Epoxy alkyl,R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or-Cl；R8、R9 Independently be-H, C1～C4Alkyl,R15～R19Independently be-H, C1～C4Alkyl ,-CF3
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 2, it is characterised in that: work as R6For R14ForTime, its structure is as shown in formula V:
Wherein, R1For-H or
R2For-H, C1～C4Alkyl,R8Substituted C3～C8Cycloalkyl,C3～C8Epoxy alkyl,N=0～4；
R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxy or halogen；
R8～R11Independently be-H, C1～C4Alkyl ,-OH,
R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen ,-CF3、-OCF3、
R20～R38Independently be-H, C1～C4Alkyl or-CF3。
3-acetenyl Pyrazolopyrimidine derivative the most according to claim 9, it is characterised in that: R1For-H or
Preferably, R2For C1～C4Alkyl,C3～C8Epoxy alkyl, N=0 or 1；R9For C1～C4Alkyl ,-OH,
It is further preferred that R2For C1～C4Alkyl,R9For C1～C4 Alkyl ,-OH or
Further preferred, R2For C1～C4Alkyl orR9For C1～C4Alkyl；
Preferably, R3～R5、R7Independently be-H, C1～C4Alkyl ,-OH or halogen；
It is further preferred that R3～R5、R7Independently be-H, C1～C4Alkyl or halogen；
Further preferred, R3～R5、R7Independently be-H, C1～C4Alkyl or-Cl；
Preferably, R15～R19Independently be-H, C1～C4Alkyl ,-OH, C1～C4Alkoxyl, halogen or-CF3；
It is further preferred that R15～R19Independently be-H, C1～C4Alkyl, halogen or-CF3；
Further preferred, R15～R19Independently be-H or-CF3；
Optimum, R1For-H；R2For C1～C4Alkyl orR3～R5、R7Independently be-H, C1～C4Alkane Base or-Cl；R9For C1～C4Alkyl；R15～R19Independently be-H or-CF3。
11.3-acetenyl Pyrazolopyrimidine derivative, it is characterised in that: its structural formula is
3-acetenyl Pyrazolopyrimidine derivative pharmaceutically acceptable salt described in 12. any one of claim 1～11.
The 3-pharmaceutically acceptable hydrate of acetenyl Pyrazolopyrimidine derivative described in 13. any one of claim 1～11.
14. 1 kinds of pharmaceutical compositions, are by the 3-acetenyl Pyrazolopyrimidine derivative described in 1～11 any one, claim Salt described in 12 or the hydrate described in claim 13 add what the pharmaceutically complementary composition of acceptable was prepared from.
3-acetenyl Pyrazolopyrimidine derivative described in 15. any one of claim 1～11, the salt described in claim 12 or The hydrate described in claim 13 purposes in preparing inhibitors of kinases.
3-acetenyl Pyrazolopyrimidine derivative described in 16. any one of claim 1～11, the salt described in claim 12 or The hydrate described in claim 13 purposes in preparation tumor.
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